PRECLINICAL TOXICITY STUDY ON

“POORA PARPAM’’

(DISSERTATION SUBJECT)

For the partial fulfillment of

Requirements to the Degree of

DOCTOR OF MEDICINE (SIDDHA)

NATIONAL INSTITUTE OF SIDDHA

CHENNAI - 600 047.

AFFILIATED TO THE TAMILNADU Dr. M.G.R MEDICAL UNIVERSITY

CHENNAI - 600 032.

DEPARTMENT OF NANJU NOOLUM MARUTHUVA NEETHI NOOLUM

APRIL 2011-2012

ACKNOWLEDGEMENT

My dissertation is soulfully dedicated to my parents Mr.M.Sivaraj, M.A,

Mrs.S.Suseela.

I express my sincere thanks to The Tamilnadu Dr. M.G.R Medical University,

Chennai-32

I express my gratitude to The Department of AYUSH, Health and Family

Welfare, Govt. of India.

I express my sincere thanks to our respectable Prof.Dr.K.Manickavasagam,

M.D(s), Director, National Institute of Siddha for granting permission and encouraging

this work.

I would express my deep sense of gratitude to our respectable

Prof.Dr.M.Murugesan,M.D(s), Dean and Head of the Department of Nanju Noolum

Maruthuva Neethi Noolum, whose excellent guidance, continuous supervision and

useful suggestions have motivated me to complete this dissertation in a good form.

I express my sincere thanks to Prof.Dr.R.S.Ramasamy, M.D (s), Head of Dept. of

Sirappu Maruthuvam for permitting me to analysis the haemotological parameters in

Clinical Pathology lab.

I express my thanks to Associate Prof.Dr.M.Rajasekaran, M.D (s), Head of Dept.

of Gunapadam for permitting me to prepare my dissertation medicine in Gunapadam

laboratory.

I express my sincere thanks to Dr.R.Madhavan, M.D(s), Dr.P.Shanmuga priya,

M.D(s), Dr.V.Manjari, M.D(s), Dr.S.Murugesan, M.D(s), Lecturers, Dept of Nanju

Noolum Marthuva Neethi Noolum, National Institute of Siddha, for their guidance,

continuous encouragement and for giving valuable suggestions to do this study.

I am very much pleased to thank Dr.V.Suba, Ph.D, Dept of Pharmacology,

National Institute of Siddha, for her memorable support and valuable guidance

throughout my dissertation work.

I express my heartfelt gratitude to Prof.Dr.Rajavelu Indra, M.D (Pathology),

Regional Institute of Ophthalmology, Govt. ophthalmology hospital, Chennai for

teaching academic and research part of my dissertation and valuable suggestion in my

carrier.

I express my thanks to Dr.A.Muthuvel, Ph.D, Dept. of Biochemistry, National

Institute of Siddha for permitting me to do the qualitative analysis of my drugs.

My sincere thanks to Mr.M.Subramaniam, M.Sc, Senior Research Officer,

National Institute of Siddha for guiding me in statistical analysis of experiments.

I am very much pleased to thank to Prof.Dr.R.Murugesan, Scientific officer,

Indian Institute of Technology, for conducting quantitative analysis of my dissertation

drug.

I express my thanks to Mrs.Sasikala ethirajalu, Asst.Director, Dept of

Pharmacognosy, Siddha Central Research Institute, Chennai, for giving authentication of

raw drug.

My sincere thanks to Mr.S.Sridhar, Mr.R.Naresh and Mrs.S.Gomathi, Lab

technician, Dept of Biochemistry and Clinical pathology for helping me in conducting

analysis of animal blood sample and qualitative analysis of drug samples.

I deeply thank all my batch mates who helped me throughout the study.

I solely and deeply regret for all the animals that lost their lives for the sake of my

study and without which I would not have been successful in my study.

CONTENTS

CHAPTERS

TITLE

PAGE NO.

I

INTRODUCTION

1

Ii AIM AND OBJECTIVES 3

III REVIEW OF LITERATURE

SIDDHA ASPECT 4

MODERN ASPECT 21

IV MATERIAL AND METHODS

4.1 COLLECTION, IDENTIFICATION PURIFICATION AND

PREPARATION OF “POORA PARPAM”

44

4.2 QUALITATIVE ANALYSIS 48

4.3 QUANTITATIVE ANALYSIS 65

4.4 TOXICOLOGICAL EVALUATION 69

V RESULTS 77

VI DISCUSSION 101

VIII SUMMERY 103

IX CONCLUSION 105

BIBLIOGRAPHY 106

ANNEXURE

P a g e | 1

CHAPTER I

1. INTRODUCTION

“The mysteries of curing and healing are hidden from the eyes, but open to the

spiritual perception of the wise” – Magnalia die

Siddha system of medicine is one of the oldest medical systems of India. The

word “Siddha” comes from the word “Siddhi” which means an object to be attained

perfection. Siddhi generally refers to Astamaa Siddhi. Those who attained or achieved

the above said powers are known as Siddhars.

Siddha system concludes body as a whole, made of five elements which are

the fundamental principles of creation, protection and destruction. With these

unparallel knowledge and uncommon development which the S iddhars had attained in

this branch of science from the very ancient times are reflected from their works as

Vatham (Alchemy), Vaidhiyam (Treatment), Yogam and Gnanam (Spirituality).

Siddha medicine means medicine that is perfect. Siddhars spend their lifetime

in experimenting the gifts of Mother Nature the herbs, the minerals and the animals.

As a result of their experiments, they could formulate so many valuable medicines

which include small herbal preparations to the potent medicines. Siddha medicine is

claimed to alleviate the root cause of the diseases by maintaining the ratio of Vatham,

Pitham and Kapham.

Siddhars are specialized in treatment of poisons. Their experience speaks from

the way of diagnosis, describing the signs and symptoms of poison in humans. The

literature survey reveals their vast knowledge in the treatment of poison that includes

metal, mineral, herbal and food poisons.

Siddhars knowledge in mentioning the specific antidote for a particular

poison is amazing by virtue of their wisdom. They also specified general antidote for

all kinds of poison. A single formulation having ability in treating all types of poisons

is their major contribution to the society.

P a g e | 2

Siddhars use metals and minerals in medicine, because of their characters like

longer shelf life, smaller the dose with greater efficacy. And the therapeutic index is

much higher compared to herbal formulations.

Siddhars obtained artificial minerals by using a special technique known as

“Vaippu Murai” and the drug acquired is known as “Vaippu saraku”. Rasam

(Mercury) is considered as the universal element present in all living matters to

certain limit. Ultimate importance is given to mercuria l salts like Pooram (Mercurous

Chloride) and Veeram (Mercuric Chloride) in medicinal formulations in treating

chronic diseases.

Siddhars knowledge about various herbs, metals and minerals are still lagging

to prove in scientific world. It is mystery to the scientific society to evaluate the exact

action of herbo - mineral formation.

According to Siddha literature Poora Parpam is given to all kinds of Vatha

disease, Gunmam (peptic ulcer), Soothaga noigal, Neerilivu (diabetes mellitus).1

In this day, arthritis is one of the lifestyle diseases commonly affecting elderly

population. The incidence of arthritis among the age group 25 to 35 years,

osteoarthritis figures as the second most prevalent disease after diabetes. And these

growing populations with arthritis are to be treated with appropriate medicine.

According to Siddha literature Poora Parpam is efficient in management of

arthritis and its related complications. In this aspect Poora Parpam may be a better

choice of drug. Since modern society is against the usage of mercurial drug as

medicine these study is more important for its own.

Here, we take the opportunity to reveal the important of this medicinal

preparation and its toxicity evaluation by animal experiment.

P a g e | 3

CHAPTER II

2. AIM AND OBJECTIVES

AIM:

To study the safety profile of the drug “POORA PARPAM”.

OBJECTIVE:

Procurement, quality assessment of the raw drugs used for the

preparation of test drug.

To evaluate the scientific rationale behind the purification process of

the ingredients of the test drug

To analyze the physical and chemical properties of the test drug.

To evaluate the toxicity profile, Acute and long-term toxicity studies

of the drug “POORAPARPAM” on animal model.

P a g e | 4

CHAPTER III

g+uk; (,urf; fw;g+uk;)

gQ;rRj tiffspy; xd;W. mWgj;J ehd;F ghlhzq;fSs;

fhzg;glhjpUe;Jk; g+uk;; ghlhz tiffSs; xd;whfNt fUjg;gLfpwJ.

,/J ,urk;> cg;G ,itfspd; $l;bdhy; nra;ag;gLfpd;w ruf;fhFk;.

itg;G Kiw

nra;Kiw:

xU ghz;lj;jpy; 16 tuhfndil (67.2 fpuhk;) fe;jfk; itj;J

cUf;fp 80 tuhfndil (336 fpuhk;) ,urk;; Nrh;j;J Johtpf;

nfhz;;bUe;jhy; fWj;Jj; J}shfptpL;k;. gpwF NtnwhU ghz;lj;jpy;

ghjpf;F nrq;fy; nghbiag; Nghl;;L mjd;Nky; miug;gb (650 kp.ypl;;;;;;;.)

fwpAg;ig itj;J cg;;gpd; Nky; Nkw;gb ,urfe;;jpia itj;J

rPiykz;; nra;;J 12 kzpNeuk; fhlhf;fpdpaha; vhpj;J vLj;;J Fsph;e;j

gpwF Nky;ghz;lj;;ij ePf;fpg; ghh;f;f g+uk; fl;baha;g; gbe;jpUf;Fk;.

Rit: cg;G> fhh;g;G.

tPhpak;: ntg;gk;.

gphpT: fhh;g;G

nra;if:

rkdfhHp>

Ngjp cz;lhf;fp>

gpj;j ePu; ngUf;fp>

cly; Njw;wp>

ckpo; ePh; ngUf;fp.

fpUkp ehrpdp.

nghJFzk;

“,ilthj #iy nahp#iy Fd;ke;

njhilthio thjkhQ; Nrhzp-apilahNjh

nthf;Fur fh;g;g+u nkhd;Nw asnthLey;

,f;Fnty;yj; NjOeh sP.”

P a g e | 5

(ngh-s;)

ey;y ,ur fh;g;g+uj;ij msTld; fUk;G nty;yj;jpy; VOehs;

nfhLf;f ,Lg;igg; gw;wpa #iy>Mq;fhq;F vhpr;riyj; jUfpd;w #iy>

thj Fd;kk;> njhilthio> thjuj;j Neha; Kjypad jPUk;.

NkYk;

“ rrptd;d fh;g;g+ uj;jpy; rhjpj;j faQ;R thrk;

grpfyp jhg Nrhgk; gTj;jpuk; gpsit Fl;lk;

trpjU fpuhzp NahL tsujp rhu Nkfk;

,rpjU kprpT #iy apitgy Nuhfk; Nghnk."

“jpuz;lth jq;Fly; thjk;

jPUQ;re; epgjpd; %d;W

kUz;l Fj;J kiuahg;G

kz;ilr; #iy fghytpb

guq;fpr; #iy gw;fpue;jp

gf;fr; #iy apit Kjy;Nghk;

,Uz;l Nkdp nghd;dpwkhk;

,JNt fw;gk; ,ak;gPNu."

G+uj;jpdhy; fak; Rthrk; gTj;jpuk; gpsit Fl;lk; fpuhzp Nkfk;

mjprhuk; ,rpT #iy Fly;thjk; rd;dp miuahg;G kz;il #iy gf;f

#iy Kjypad jPUk;

g+uk; - Ruk;> kQ;rl; fhkhiy> gpj;j Njhlk;> rPPPjNgjp> ePh;f;Nfhit>

tpuzre;ep> Mwhj tpuzq;fs;> Nkf tpahjp> nrhpahik> the;jp> Ngjp>

fpUkp Neha;> fPPy;thjk;> nrhwp> rpuq;F> kyge;jk; Kjypaitfl;Fk;>

jiytyp Nghd;w kw;iwa Neha;fSf;F cgNahfpf;fgLfpwJ.

g+u Rj;jp

g+uj;jpYs;s kypdq;fis ePf;Ftjw;Fk;> tplf;Fwp Fzq;fs;

Vw;glhjpUg;gjw;Fk; mjpf tPhpaj;ijf; nfhLj;jw;Fk; ,jid Rj;jp

nra;a Ntz;Lk;.

P a g e | 6

Rj;jp Kiwfs;:

1. fk;khW ntw;wpiy>kpsF Mfpa ,uz;ilAk; fhy;gyk;(8.75 fp;)

tPjk; epWj;njLj;J rpwpJ ePh; tpl;L miuj;J> fy;fj;ij xU gb(1.3 ypl;)

ePhpy; fye;J> xU gyk; (35 fpuhk;) g+uj;ij rPiyapy; Kbe;J

Jyhae;jpukha; ePhpy; mkpOk;gb nra;J> rpW jPahy; vhpf;f Ntz;Lk;.

ePh; Kf;fhw;gq;F Rz;ba gpwF> g+uj;ij vLj;J ePh;tpl;Lf; fOtp

nta;apypy; cyh;j;jp vLf;f Rj;jpahk;.

2. xU gyk; (35 fpuhk;) g+uj;jpw;F Kiyg;ghypdhy; %d;W kzpNeuk ;

RUf;Ff; nfhLj;Jg; gpwF nts;isg;g+z;Lj; ijyj;jpdhy; xd;gJ

kzpNeuk; RUf;fpl;L vLj;Jf;nfhs;sTk;.

,Nyfpaq;fspy; Nrh;f;f g+uj;ij> KRKRf;ifr; rhw;wpdhy;

RUf;fpl;Lf; Rj;jp nra;aTk;.

kUj;Jt gad;fs;1

g+u gw;gk; ;

Rj;jp nra;j g+uj;ijf; fy;tj;jpypl;L VOehs; ed;wha; miuj;J

gpwF Nrfhpj;Jf; nfhs;sTk;. ,JNt g+u gw;gnkd;W ifahsg;gLfpd;wJ.

msT:

miu cSe;njil (32 kp.fp;uhk;) apypUe;J %d;W cSe;njil

(195 kp.fp;uhk;) tiu cgNahfpf;fyhk;. ,uz;L cSe;njil (32kp.fp;uhk;)

f;F Nkw;glf; nfhLj;jhy; NgjpahFk;.

ehs; msT : VOehs;

JizkUe;J: fUk;G nty;yk;.

jPPUk; Neha;fs:

Ruk;> kQ;rl;fhkhiy> gpj;jNjhlk;> rPPPjNgjp> ePh;f;Nfhit>

tpuzre;jp> Mwhj tpuzq;fs;> Nkf tpahjp> nrhpahik> the;jp> Ngjp>

fpUkpNeha;> fPPy;thjk;> nrhwp rpuq;F> kyge;jk;> jiytyp Kjypaitfs;

jPUk;.

P a g e | 7

G+u cg;G5

itag;gh gl;re;jhd; nrd;wgpd;G

tifahf naLj;Jg;ghh; cg;GkhFk;

nra;ag;gh Rz;zhk;Gf; Fifapypl;Lr;

rpwg;ghf Nky;%br; rPiy nra;J

ieag;gh gj;njUtpw; Glj;ijg;NghL

eykhFk; eykhFk; g+ur;Rz;zk;

ifag;gh gl;Lnjd;why; nte;JePWk;

fdpthf apd;dnkhU fzf;Ff;NfNs.

fzf;fhf Kd;nrhd;;d nraePh;jd;dpy;

fhhpakha; eidj;JNk nrhy;yf;NfS

gpzf;fhf mQ;RRz;zf; Fifapy; itj;Jg;

NgrhNj Glk;Nghl;L naLj;Jg;ghU

jzf;fkha;f; fLQ;Rz;zk; g+ur;Rz;zk;

jhl;bkhfnahUtUld; nrhy;yNtz;lhk;

tzf;fkha;r; n[frhy tpj;ijahLk;

thjkil jpwg;ghFk; tz;ikahNk.

tz;ikah apr;Rz;ze; jd;dpw;whDk;

tbthfr; n[frhy klq;fpg;Nghr;R

jd;ikaha; Rz;znky;yh kpjd;fPohr;R

jathff; fiuapy;iy fzf;Fkpy;iy

cz;ikaha; xg;gidjhd; nrhy;yntd;why;

cWjpaha; KbahJ cz;iknrhd;Ndd;

ntz;ikaha; tpk;kptpk;kp aiyaNtz;lhk;

ntUshNj gPq;fhdpy; gjdk;gz;Nz.

- ahNfhG Rz;zf;fhz;lk; - 18>19>20

ghly;fs; : 38-40.

P a g e | 8

nghUs;:

G+uk; xU gyk; vLj;Jf; nfhz;L fLq;fhur; nraePu; tpl;L

miuj;J gPq;fhdpy; itj;J gjpide;J ehl;fs; nrd;w gpd;G vLj;Jg;

ghu;f;f cg;ghf ,Uf;Fk;. ,ij Rz;zhk;G Fifapy; Nghl;L %b rPiy

kz; nra;J gj;J vUtpy; Glj;ijg; NghlTk;. Mwpa gpd; vLj;Jg;

ghh;f;f g+uk; Rz;zkhfp ,Uf;Fk;. ,e;j Rz;zj;ij Nkw;gb nraePupy;

eidj;J vLj;J gQ;R Rz;zf; Fifapdpy; itj;J rpy;Y %b rPiy

kz; nra;J Glk; Nghl fLikahd Rz;zkhFk;. gpd;G ,r;Rz;zj;ij

gPq;fhdpy; gjdk; nra;J itf;fTk;. ,jdhy; vy;yh KiwfSk;

n[akhFk;. urid vd;w kil ,jdhy; jpwe;J NghFk;. ,e;j

Rz;;zj;jpy; vy;yh tpj;ijfSk; mlf;fk;. kw;w vy;yh Rz;zq;fisAk;

tpl ,J NkyhdJ.

,urf;fw;g+u itg;G fw;gk; ;13

cfe;Jnfhz;L epw;gjw;F urfw;g+u

KWjpAld; nrhy;YfpNwd; Gyj;jpahNfs;

tFe;Jepd;w jphpfLF nte;jaKnkhd;wha;

khh;f;fKld; tif%d;W tuhfd; J}f;fp

gfh;e;Jepd;w kj;jQ;rhh; tpl;liue;J

ghyfNd FifNghy gz;zpf;nfhz;L

Rfe;jKld; urNfj;jp ypq;f%d;WQ;

Rj;jKld; nghbg;gz;zp Fifapy;NghNl.

Nghlg;gh rl;bKf;fh Yg;igf;nfhl;b

Gj;jpAl df;Fifia gjpaitj;J

ehlg;gh Nky;rl;b nfhz;L%b

eykhf kz;rPiy MWrhkk;

Jg;Gutha; jPnahpe;J naLj;Jg;ghh;j;jhy;

%h;f;fKld; urNkwp gw;gkhr;Nr.

Mr;rg;gh gw;gnkd;w urfw;g+uk;

mg;gNd gf;Ftkh naLj;Jf;nfhz;L

tpr;rg;gh fpue;jpKjy; Fl;lj;Jf;Fk;

tpirahd gTj;jpuKQ; #iyf;nfy;yhk;

P a g e | 9

ghr;rg;gh gztpilfhy; rh;f;fiuapy; jhDk;

gwf;Fklh rfytp\ Nuhfnky;yhq;

fhr;rg;gh g+unkd;why; urfw;g+uk;

fz;lTld; jPUklh fUtha;g;ghNu

ghug;gh apd;dnkhU #l;rq;NfS

gTj;jpu FNlhhpnad;w %ye;jd;id

fhug;gh aPUs;sp Jk;igNtis

fLfNt jl;baij jl;bdf;fhy;

Ntug;gh ma;j;jJTk; Mwpg;NghFk;

tpirahd kUe;jpJjhd; gprNfhapy;iy

Mug;gh mwpthh;f spe;j%yk;

mUikntF mUikalh mwpe;JghNu.

- mf];jpah; G+uz fhtpak; Mapuk; - 236>237

nghUs;:

jphpfLF> nte;jak; tiff;F %d;W tuhfd; vLj;J Ckj;ij

rhW tpl;liuj;J FifNghy; nra;J nfhs;s Ntz;Lk;. ,urk;> fe;jfk;>

,ypq;fk; %d;iwAk; nghb nra;J ,f;Fifapypl;L xU rl;bapy;

Kf;fhy; ghfk; tiu cg;G nfhl;b mjpy; ,f;Fifia itj;J rl;b

tha;f;F nghUj;jkhd Nkw;rl;b nfhz;L %b kz;rPiy nra;J

cyHj;j Ntz;Lk;. cyHj;j gpd;G ,jid MWrhkk; jPapl;L vhp;j;Jg;

ghu;f;f ,urfw;G+u gw;gk; Nky; rl;bapy; gbe;jpUf;Fk;. ,jid Ruz;b

vLj;J gj;jpug;gLj;j Ntz;Lk;. ,JNt fw;g kUe;jhFk;.

msT: fhy; gzNtil (122 kp.fp.)

JizkUe;J: rHf;fiu

jPPUk; Neha;fs;:

fpue;jp> Fl;lk;> gTj;jpuk;;> #iy> rfy tplk;.

P a g e | 10

G+uk; NrUk; gpw kUe;Jfs;4

1. ,urfw;g+uty;yhjp

msT: jhd;wpasT 2 Ntis

jPUk; Neha;fs;: thjk;> #iy> jpkpu;> iffhy; tPf;fk>; cisr;ry;> Gw;W>

kU> Fd;kk; jPUk.;

2. G+uf;fl;L

msT: muprp vil

mDghdk;: gidnty;yk;> ntz;nza;> ghNyL KjypaitfshFk;.

jPUk; Neha;fs;: thANeha;> thj#iy> Fd;kNeha;> njhilthio>

,uzq;fs; Kjypa Neha;fSk; jPUk;.

3. ,urf;fw;g+unkOF

msT: 1 Kjy; 2; Fd;wp vil

mDghdk;: gidnty;yk>; ey;ynty;yk;> ntz;nza;> ghNyL.

jPUk; Neha;fs;: guq;fpg;Gz;> NkfNeha;> #iyNeha; Mfpaitfs; jPUk;.

%d;Wehs; MWNtis nfhLj;jhy; NghJkhdJ.

4. ,urf;fw;g+unkOF

msT: 1 Kjy; 2; Fd;wp vil

mDghdk;: nty;yk;> Rf;if miuj;j tpOJ> ntz;nza; NtW

,sf tiffs; Kjypadthk;.

jPUk; Neha;fs;: kyf;fl;L> Fd;kNeha;> gPypif> fpue;jp;> ,uzq;fs;> Nkf

Neha;fs; Kjypaitfnsy;yhk; jPUk;.

5. NfhNuhrid nkOF

msT: 1 Kjy; 2; Fd;wp vil

mDghdk;: rPdpr;ru;f;fiu> gidnty;yk;.

jPUk; Neha;fs;: #iy Neha;fs>; miuahg;G> fz;lkhiy> #jfthA>

Nkf ,uzq;fs; jPUk.;

6. ntl;il nkOF

msT: 1 Fd;wp vil

mDghdk;: gidnty;yk;;> ntz;nza;> ghNyL.

jPUk; Neha;fs;: ntl;il NehahdJ vj;jid ehs; gl;ljhapDk; jPu;e;J

tpLk;. Mz;FwpapYs;s Gz;> ePu;j;jhiuapYs;s Gz;;

jPUk.;

P a g e | 11

7. g+u khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

mDghdk;: jha;g;ghy;> tpsf;nfz;nza;> gidnty;yk;> Rf;F

FbePu;> Nrhk;Gf; FbePu.;

jPUk; Neha;fs;: thA Neha;fs;> kyge;jj;jpdhYz;lhd Neha;fs;>

gps;isfSf;Fz;lhFk; typg;G> #iy Neha; jPUk;.

8. thA khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

mDghdk;: gidnty;yk;> Rf;if miuj;j tpOJ.

jPUk; Neha;fs;: thA Neha;fs;> #iy> gps;isfSf;Fz;lhFk; typg;G

jPUk;

9. et];g+u khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

mDghdk;: jha;g;ghy;> gidnty;yk;> ntz;nza;> ghNyL>

rPdpr;ru;f;fiu> thiog;gok; Kjypadthk;.

jPUk; Neha;fs;: khe;jk;> Ruk;> ,rpT> thA #iy Neha; jPUk.;

10. jPu; me;jh];fhd; khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

mDghdk;: gidnty;yk;> ey;ynty;yk;> thiog;gok;> Rf;if

miuj;j tpOJ.

jPUk; Neha;fs;: mz;lthjk;> neupf;fl;Lr;Ruk;> fPy;thA> thj Neha;fs;>

Fd;kNeha; fl;b Kjypaitfs; jPUk;.

11. fw;g+u rpe;jhkzp khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

mDghdk;: ru;f;fiu> nty;yk;> nea;.

jPUk; Neha;fs;: vy;yhtif RuNeha;fSk; jPUk;. fPy;thA> Klf;FthA>

#iyNeha; KjypaitfSk;; jPUk;.

12. #iy,uhl;r khj;jpiu

msT: 1 Kjy; 2 khj;jpiu

fhiy khiy ,uz;L NtisAk; cgNahfpf;fTk;. VO

ehl;fs;.

mDghdk:; gidnty;yk;;> ntz;nza;> ghNyL> Rf;if miuj;j

tpOJ.

jPUk; Neha;fs;: fpue;jp Neha;> #iy Neha;> #iyf; fl;L;> gpuNkfk;

Fd;kNeha;> thA Neha; Kjypa Neha;fs; jPUk;.

P a g e | 12

13. `y;jPg+u khj;jpiu

msT: 1 Kjy; 2 khj;jpiu.

mDghdk;: gidnty;yk;;> ru;f;fiu> jha;g;ghy; Kjypadthk;.

jPUk; Neha;fs;: vy;yhtif RuNeha;fspYk; cgNahfpf;fyhk;.

14. Rff; fopr;ry; khj;jpiu

msT: 1 Kjy; 2 khj;jpiu.

gLf;iff;Fg;NghFk; NghJ xU khj;jpiuia thapy;

Nghl;Lf;nfhz;L nfhQ;rk; nte;ePu; rhg;gpl;Lg; gLj;Jf;

nfhs;sTk;. fhiyapy; nrk;ikahf fopr;ry; MFk;.

jPUk; Neha;fs;: clypYs;s thj> gpj;j> fg jPa ePu;fs; ntspg;gl;L

cly; MNuhf;fpaKz;lhFk.;

15. ghy rQ;rPtp khj;jpiu

msT: 1 Kjy; 2 khj;jpiu.

mDghdk;: jha;g;ghy;> Njd; Kjypadthk;

jPUk; Neha;fs;: Foe;ijfspd; vy;yh Neha;fSf;Fk; nfhLf;fyhk;.

Nehapw;Fj; jFe;jgb %d;W ehs; Kjy; VO ehs;

cgNahfpf;fyhk;.

16. xsitghy rQ;rPtp khj;jpiu

msT: 1 Kjy; 2 khj;jpiu.

mDghdk;: jha;g;ghy;> nghLjiyf; FbePu;> Ezh ,iyf; FbePu;

Kjypadthk;

jPUk; Neha;fs;: Foe;ijfspd; vy;yh Neha;fSf;Fk; nfhLf;fyhk;.

17. ,ur fw;g+u gw;gk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza;> ,sf tiffs;

Kjypadthk;.

jPUk; Neha;fs;: thA Neha;fs;> Ruk;> rd;dp jPUk.;

18. g+u rl;ru gw;gk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza;> thiog;gok;> Rf;Ff; FbePu;>

Rf;if miuj;j tpOJ Kjypadthk;

jPUk; Neha;fs;: Ruk;> rd;dp> #iy> Fd;kk;> thA Kjypaitfs; jPUk;.

P a g e | 13

19. Kbg+u gw;gk;

msT: 1 Kjy; muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza; Kjypadthk;.

jPUk; Neha;fs;;: Gz;fs;> fl;bfs;> ntl;il> #iy> Nkfk; jPUk.;

20. m\;l fw;g+uk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza; Kjypadthk;

jPUk; Neha;fs;: tapw;Wtyp> Fd;kNeha;> Ruk;> rd;dp> ,rpT> typg;G.

21. ,urfw;g+ur; nre;J}uk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza; Kjypadthk;

jPUk; Neha;fs;: Ruk;> rd;dp> Fd;kNeha;> #iyf;fl;L jPUk.;

22. g+u gjq;fk;

msT: 1 Kjy; 2 muprp vil(65 – 130 kp.fp)

mDghdk;: Vyf;fha; J}s; tuhfndil> ntz;nza; tuhfndil

,t;tpuz;ilAk; Nru;j;J nfhs;s Ntz;Lk;.

jPUk; Neha;fs;: fpue;jp Neha;> Gz;> Giu jPUk.;

23. rz;lkhUj nre;J}uk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> ntz;nza;> ,sf tiffs;> Rf;if

miuj;j tpOJ Kjypadthk;.

jPUk; Neha;fs:;; Ruk;> rd;dp> Fd;kNeha;> #iyf;fl;L jPUk.;

24. Kg;g+ur; nre;J}uk;

msT: 1 Kjy; 2 muprp vil. (65 – 130 kp.fp)

mDghdk;: gidnty;yk;;> jpupgiy ,sfk;> jpupfLF ,sfk;

Kjypadthk;.

jPUk; Neha;fs;: Ruq;fs;> rd;dpfs;> tapw;Wtyp> Fd;kNeha;> #iy Neha;

#jfthA> NkfNeha; Mfpaitfs; jPUk.;

- mDNghf itj;jpa etePjk; - ghfk; 4

P a g e | 14

Gw kUe;Jfs; 1

G+uf; fspk;G

cgNahfk;: ,ijg; gwq;fpg; Gz;Zf;F Nky; NghlTk;.

G+u vz;nza;

msT: miu (14 kp.ypl;) Kjy; xU mTd;]; (28 kp.ypl;)

%d;W ehs; fhiy khj;jpuk; nfhLf;fTk;.

gj;jpak;: cg;G> Gsp ePf;fTk;.

cgNahfk;:

kyk; fopAk;> Foe;ijfspd; khe;j Neha; jPUk;.

Nkf Nuhfq;fSk; Fd;kKk; ePq;Fk;. ,t;ntz;nzAld;

tha;tplq;fj;J}s; Nrh;j;J cgNahfpf;f fpUkp ePq;Fk;.

G+ug; nghb

cgNahfk;:

nfhWf;Ff;F (Mz; Fwpapy; fpue;jpapdhy; cz;lhFk; Gz;)

,j;J}is NkNy J}tptuf; FzkilAk;. ,g;nghbia tl

E}yhh; “gwq;fpf; Nfrhp” vd;W $Wth;.

P a g e | 15

G+u eQ;R

G+u eQ;Rf; FwpFzk;

G+uk; tplkpj;jpUe;jhy;>

Kfj;jpy; nrt;thg;Gg; Nghy Kfg;gU>

Nth;f;FU mjpfkha; cz;lhjy;>

khh;gpd; gs;sj;jpy; gUf;fl;bg; Gz;zpy; uzk; fhZjy;>

gf;fj;jpy; typ>

thapy; fhuk; glhjgb Gz;zhjy;>

gPrtPf;fk;>

cz;zhf;fpy; tpuzk;>

Ngjp> ,uj;jf; fopr;ry; Kjypa Jh;f; Fzq;fisf;

fhz;gpf;Fk;.

G+u eQ;R KwpT

epyg;gidf; fpoq;F>

ty;yhiu Nth;>

nghd;dhq;fz;zp Nth;>

fz;L ghuq;fp>

Mfpa ,it xt;nthd;Wk; jdpj;jdpf; fhw;gyk; (8.75 fpuhk;)

vLj;J> xd;W Nrh;j;Jf; FbePhpl;L> ,UNtis tPjk; ,uz;L my;yJ

%d;W thuk; gj;jpaj;Jld; mUe;j eQ;R ePq;Fk;.

- Fzghlk; jhJ rPt tFg;G 282 - 288

P a g e | 16

vUf;F

NtW ngau;:

mUf;fd;

gad;gLk; cWg;G:

,iy> g+> gl;il> Ntu;.

Rit: ifg;G> fhuk;> kJuk;.

jd;ik: ntg;gk;

gpupT: fhu;g;G

nra;if:

vUf;fk;ghYf;F - ehrpePu;tu;j;jpdp

vUf;fk;ghy;

vUf;fk;ghw; fl;bfis Nafiuf;Fk; tha;itj;

jpUf;fwNt nfhd;WtpLe; jPuhr; nrUf;fhd

re;jp typjPu;f;FQ; rhu;e;jgy rpe;J}uk;

Kd;d Kbf;Fnkd NthJ.

nts;nsUf;fk; ghy;

typapd; typfSf;F khthj rd;dp

vypapd; tplQ;Ruq;f nsy;yhk; typafw;Wf;

fhiyj; njhoNj flw;NrUk; nts;nsUf;fk;

ghiyj; njhLthu;f;Fg; ghu;.

P a g e | 17

cgNahfpf;Fk; Kiw3

ghiy RSf;F> fl;b> ntl;ilahYz;lhd fPy;thA>

eupj;jiy thA ,itfSf;Fg; gw;wplyhk;.

gy; Neha;> gy; nrhj;ij> gy; <Wfl;L ,itfSf;Fg;

ghiyj; njhl;Litf;fyhk;.

ghiy cyu;j;jp 65 my;yJ 130 kpfp gidnty;yj;jpy;

itj;Jf; nfhLj;J cg;gpy;yhg; gj;jpak; itf;f vypf;fb

eQ;R ePq;Fk;.

ghiy Kjd;ikahf nfhz;L nra;ag;gLfpd;w

“mu;f;f\Puj;ijyk;” kfhthj Neha;fisf; fz;bf;Fk;.

,g;ghy; mz;l mUf;fd; nra;ePu;; nra;a cjTk;.

xU Njf;fuz;b msT ey;nyz;nzapy; 7 Jsp

vUf;fk;ghiy tpl;L ed;wha;f; FYf;fp ehrpf;Fs; 2-3 Jsp

tpl msTfle;j Jk;ky; cz;lhFk;. rpurpYz;lhd

ePiunay;;yhk; ntspg;gLj;Jk;.

fhf;if typf;Fr; rpfpr;ir nra;Ak; NghJ Kjypy;

,r;rpfpr;ir nra;tjpdhy; %isia mDrupj;j rPjsj;ij

mfw;Wk; me;jj; Jk;kiy epWj;jNtz;Lkhapd; Kfj;jpy;

Fspue;j ryj;jhy; mbj;Jf; Fspu;e;j ryj;ijf; nfhz;L

ehrpia Rj;jg;gLj;j Ntz;Lk;.

vUf;fk;ghiyf; fPy;gpbg;G> tPf;fk; ,itfl;F Nkw;wltp

mbapypUf;Fk; kz;iz Nkw;W}tpf; fhatpl FzkhFk;.

tp\ fbthapy; vUf;fk;ghiyj; jltpg; Gz;zhf;f> tp\k;

Kwpe;J Nghk;.

,d;Dk; ,e;j ghYld; ,ju ruf;Ffisf; $l;bj; je;j

Nuhfq;fSf;Fk;> thjNuhfq;fSf;Fk; mtpo;jq;fs;

nra;tJz;L.

P a g e | 18

tr;rpuje;j nkOF11

,uz;nlhU rpl;bif vLj;Jj; je;j Rj;jp nra;Jtug;

gy;typ> gy;nrhj;ij> gy;rpYrpYg;G Kjypaitfs; Nghk;.

vUf;fk; ghy; ijyk;

cr;rpapy; rpwpJ Nja;j;Jg; gpd;G Njfnkq;Fk; Nja;j;J

KWf;fp gpbf;f ru;tNjftyp> Filr;ry;> ,rpT Kjypaitfs;

Nghk;.

eQ;Rf; FwpFzk;6

jtwhf cgNahfpj;Jtpl;lhy; me;j ,lj;jpy; Gz;iz

cz;L gz;Zk;. cs;Sf;Ff; nfhLj;J tpl;lhy; tha;> tapW

Kjypait ,uzkhtNjhL> the;jp> Ngjp KjypaitfisAk;

cz;lhf;Fk;.

KwpT

1. ru;f;fiuAk;> 4 fpuhk; vs;isAk; Nru;j;J cz;z Ntz;Lk;.

2. gUj;jpapiyiaf; fhbtpl;L miuj;J xU ghf;fsT

cl;nfhs;s Ntz;Lk;.

3. kQ;rl;nfhbAk;> rpw;whkzf;F nea;Ak; fye;J

cr;rpfuz;basT nfhLf;f Ntz;Lk.;

4. xU Ntisf;F 20 kpyp ey;nyz;nza; tPjk; %d;W my;yJ

Ie;J Ntis cl;nfhs;s Ntz;Lk;.

5. 10 fpuhk; khtpyq;fg;gl;iliaj; jz;zPu; tpl;liuj;J

cl;nfhs;sTk;.

P a g e | 19

ntq;fhak;

,/J ,e;jpah KOtJk; gapuplg;gLfpwJ. ,jd; ,iy jhnsdg;

ngau; ngWk;. ,j;jhl;fSk; fpoq;Fk; czT tiffspYk;> fpoq;Fk;

tpijAk; kUe;J tiffspYk; gad;gLfpwJ.

NtW ngau;:

<Us;sp>;; cs;sp> <uTs;sp> <untq;fhak;> fhak;> Rf;fpue;jk>;

epr;rpak; gyhz;L.

Rit: fhu;g;G

jd;ik: ntg;gk;

gpupT: fhu;g;G

nra;if:- ntg;gKz;lhf;fp

rpWePug;ngUf;fp

Nfhioafw;wp

#jfKz;lhf;fp

jbg;Gz;lhf;fp

cs;soyhw;wp

fhkk;ngUf;fp

nghJf;Fzk;:

ntg;g% yq;fpue;jp tPWuj;j gpj;jKld;

nrg;Geh mf;fue;jP uhj;jhfk; ntg;Gf;

fLg;GWke; jQ;re;ep fhrk;tapw;W g;gy;

jbg;NgWk; ntq;fhaj;jhy;.

– mfj;jpaH Fzthflk;.

ntq;fhaj;jhy; clypd; ntg;gk;> %yk;> rpuq;F> FUjpaoy; Neha;>

mf;fuk;> ePu;Ntl;if> fopr;ry; ,it ePq;Fk;. Mdhy; ke;jk;> Kg;gpzp>

,Uky;> tapw;Wg;gprk; ngUFk;.

P a g e | 20

gr;ir fpoq;ifj; jpd;d mJ ngz;fSf;Fr; #jfj; jiliag; Nghf;fp>

FUjp rpf;fiy mWf;Fk;> ePiu ngUf;Fk;.

ntq;fhaj;Jld; 2-3 kpsF Nru;j;Jz;z espu;r;Ruk; jzpAk.;

ntq;fhaj;ij cg;Gld; $l;bAz;z tapw;Wtyp> FUjpaoy; Neha;

NghFk.;

FbePupl;L Fbf;f ePu;f;fl;L> ePnuupr;ry;> ePu;f;fLg;G ePq;Fk.;

nea; tpl;L tjf;fpAz;z clydy; jzpAk;.

Fk;gp rhk;gypy; Ntftpl;liuj;J fiuahf; fl;bfs; kPJ itj;Jf;

fl;l> fl;biag; gOf;fitj;J cilf;Fk;.

fpoq;fpd; rhw;iw Kfu %u;r;irAk;> #jpfhrd;dpAk; jzpAk.;

fhjpy; 2-3 Jsptpl fhJ Neha; ePq;Fk;.

Nky; g+r g+r;rpfbfs;> Njs; nfhl;ly; ,itfshYz;lhFk; typ> Gil

Kjypa Neha;fs; NghFk;.

fhbAld; fye;J cl;nfhs;sj; njhz;ilg; Gz; khWk;.

fhbAld; Xu; msT $l;bf; fha;r;rpf; nfhLf;f fhkhiy> tapw;Wtyp>

<uy; tsu;r;rp xopAk;.

fLnfz;nzAld; XusT fye;J g+r fPy;thjk; Fzg;gLk;.

jdpr;rhw;iw cl;nfhs;s Gifapiy eQ;R khWk;.

P a g e | 21

MERCUROUS CHLORIDE (CALOMEL)

INTRODUCTION

Mercurous chloride, mercury (I) chloride or calomel is a white

crystalline powder and very slightly soluble in water. Mercurous chloride is a less

dangerous poison than mercuric chloride because it is much less soluble; but it is

highly toxic if retained in the body. It is also found in nature as horn quicksilver. It

was used medicinally as a purgative, cathartic, liver stimulant, and to eliminate

parasitic worms, but is rarely so used today because it is readily decomposed into

metallic mercury and the very poisonous mercuric chloride found on exposure to

sunlight or if heated in the presence of moisture.

COMPOSITION Mercurous chloride (85.0% Hg, 15.0% Cl).

TESTS Completely volatilizes on charcoal, without melting.

DISTINGUISHING CHARACTERISTICS

The sectile character and the adamantine luster distinguish it from

everything except from the silver halides. Silver minerals melt but do not volatilize

completely on the charcoal, leaving a flattened silver residue. In a mercury association

ore the fluorescence is significant.

CRYSTAL DESCRIPTION - Tetragonal -- Ditetragonal Bipyramidal

Usually found as crystals, often minute and coating other minerals.

Most often tabular, sometimes pyramidal. Commonly in skeletal parallel growths

rather than good individual crystals.

P a g e | 22

GENERAL PROPERTIES

Chemical formula: Hg2Cl2

Chemical name: Mercurous Chloride

Mineralogical name: CALOMEL

Color: White, Yellowish gray, Gray, Yellowish white, Brown

(Darkening on exposure to light).

Habit: Earthy - Dull, clay- like texture with no visible crystalline

affinities.

Prismatic - Crystals Shaped like Slender Prisms (e.g.

tourmaline).

Molar mass: 472.09 g/mol

Solubility in water: 0.2 mg/100 ml.

Solubility: Insoluble in ethanol, ether

Luminescence: Fluorescent, Short UV=dark red, Long UV=dark red.

Luster: Adamantine - Resinous, translucent; fluorescent red.

Streak: pale yellowish white

Hardness: 1.5-2 - Talc-Gypsum

Specific gravity: 6.5

Density: 6.4 - 6.5, Average = 6.45

Melting point: 525 °C (triple point)

Boiling point: 383 °C (sublimes)

Diaphaneity: Translucent to sub translucent

Fracture: Sectile - Curved shavings or scrapings produced by a knife

blade, fracture conchoidal (e.g. graphite)

Refractive index (nD) - 1.973

Highly reactive with metals except Sn, Pb, Ag, Au

P a g e | 23

STRUCTURAL PROPERTIES33

Mercury is unique among the group of 12 metals for its ability to form the M –

M bond so readily. Hg2Cl2 is a linear molecule. The crystal structure is unit cell

distorted octahedral coordination of mercury. The chemical bonding are

The Hg–Hg bond length of 253 pm (Hg – Hg in the metal is 300 pm) and the

Hg – Cl bond length in the linear Hg2Cl2 unit is 243 pm.

The overall coordination of each Hg atom is octahedral as, in addition to the

two nearest neighbors, there are four other Cl atoms at 321 pm.

PREPARATION

Mercurous chloride is prepared by sublimation from a mixture of mercury and

mercuric chloride or by precipitation from a mercurous chloride solution on adding

chloride ion. . It may be obtained by heating mercury in chlorine, or by reducing

mercuric chloride (corrosive sublimate) with mercury or sulphuric acid. It is

manufactured by heating a mixture of mercurous sulphate and common salt in iron

retorts, and condensing the sublimed calomel in brick chambers. In the wet way it is

obtained by precipitating a mercurous salt with hydrochloric acid. Long continued

boiling with water gives mercury and mercuric chloride; dilute hydrochloric acid or

solutions of alkaline chlorides convert it into mercuric chloride on long boiling.

CHEMICAL REACTIONS

Mercurous chloride forms by the reaction of elemental mercury and mercuric chloride

Hg + HgCl2 → Hg2Cl2

It can be prepared via metathesis reaction involving aqueous mercury (I) nitrate using

various chloride sources including NaCl or HCl.

2HCl + Hg2(NO3)2 → Hg2Cl2 + 2HNO3

P a g e | 24

Ammonia causes Hg2Cl2 to disproportionate:

Hg2Cl2 + 2NH3 → Hg + Hg (NH2) Cl + NH4Cl

REACTIVITY PROFILE

Mercurous Chloride is incompatible with acetylene, ammonia, chlorine

dioxide, azides, calcium (amalgam formation), sodium carbide, lithium rubidium and

copper.

HEALTH HAZARD25

Acute poisoning can result from inhaling dust concentrations of 1.2-8.5

mg/m 3 in air; symptoms include pain and tightness in chest, coughing, and difficulty

in breathing. Compound is an irritant, cathartic, or purgative; rarely, “calomel

sickness,” a benign reaction with fever and rash, appears after about 1 week; seldom

causes systemic poisoning but may be fatal if retained to 30-40 mg/kg. Contact with

eyes causes mild irritation.

EMERGENCY25

Eye contact: Immediately flush the eye with water. If irritation persists, go for

medical help.

Skin contact: Wash off with soap and water.

Do not discard into a sink or into normal solid waste containers.

Wear Protective equipment, safety glasses, and gloves.

P a g e | 25

TOXICOLOGICAL ASPECT35

May be fatal if swallowed or inhaled. Chronic exposure may lead to

systemic effects and build-up of mercury in the brain, liver and kidneys. May cause

memory loss, tremors and other serious effects.

LD50 - 210 mg/ kg.b.wt on oral administration to rat. 35

REPRODUCTIVE AND

DEVELOPMENTAL TOXICITY

CALOMEL

MERCURY

CA Prop 65 Developmental Toxin

Yes

Yes

Symptoms of Calomel Exposure from the International Chemical Safety Cards

(ICSC) 35

ROUTE OF EXPOS URE

SYMPTOMS

FIRST AID

Inhalation

Cough. Sore throat.

Fresh air rest. Refer for

medical attention.

Skin

May be absorbed. Redness

may develop.

Remove contaminated

clothes. Rinse skin with

plenty of water or shower.

Refer for medical

attention.

P a g e | 26

Eyes

Redness may be developed.

First rinses with plenty of

water for several minutes

(remove contact lenses if

easily possible) then take

to a doctor.

Ingestion

Abdominal pain. Diarrhoea.

Vomiting. Metallic taste.

Rinse mouth. Induce

vomiting (ONLY IN

CONSCIOUS PERSONS).

Refer for medical

attention.

AQUATIC ECOTOXICITY35

All Toxic Effects for Organism Group

ORGANISM GROUP

EFFECTS NOTED

Aquatic Plants Cell(s)

Echinoderms Development

Fish Mortality

Insects Intoxication

Mollusks Accumulation, Biochemistry

Nematodes and Flatworms Development, Mortality

Zooplankton Mortality

P a g e | 27

ORGANISM GROUP

AVERAGE ACUTE TOXICITY

Fish

Moderately Toxic

Nematodes and Flatworms

Slightly Toxic

Zooplankton

Very Highly Toxic

P a g e | 28

CALOTROPIS GIGANTEA (ERUKKU)

TAXONOMICAL CLASSIFICATION

Kingdom: Plantae

Division: Magnoliophyta

Class: Magnoliopsida

Sub-class: Asteridie

Order: Gentinales

Family: Asclepiadaceae

Genus: Calotropis

Species: gigantea

SYNONYMS

Sanskrit: Arka, Alarka

English: Gigantic swallow wort, Mudar, Crown flower, crown plant,

Giant milkweed, rubber bush, bow-string hemp.

Hindi: Madar

Kannada: Ekkemale

Telugu: Mandaramu, Ekke, Jilledu, Arka

Malayalam: Errikka

ORIGIN AND GEOGRAPHIC DISTRIBUTION

Calotropis gigantea is native to continental Asia and South-East Asia and has

been introduced in the Pacific Islands, Australia, Central and northern South America

and Africa. Used as an ornamental near villages, temples and as a weed.

P a g e | 29

DESCRIPTION

Large shrub or small tree up to 4m tall, much-branched at base, stems erect,

up to 20 cm in diameter; bark pale grey, longitudinally cracked; young shoots

woolly hairy; latex in all parts.

Leaves opposite, decussate, simple and entire, sessile; stipules absent; blade

broadly ovate to oblong to obovate, 9.5 to 20 cm × 6 to 12.5 cm, base cordate

with semi-amplexicaul lobes, apex almost acute, short-hairy beneath.

Inflorescence an axillary, umbellate to almost corymbose cyme up to 12.5 cm

in diameter, peduncle 6 to 12 cm long, stout, secondary branches up to 2 cm

long.

Flowers bisexual, regular, 5-merous, white, cream, lilac or purple; pedicel 2.5

to 4 cm long, densely woolly hairy; calyx lobes broadly ovate, ovary superior,

2-celled, gynostegium up to 1 cm long, stigma head star shaped.

Fruit a pair of follicles, each follicle ovoid, boat-shaped, inflated, 6.5 to 10 cm

× 3 to 5 cm, many seeded. Seeds ovoid, 5 to 6 mm long, with 2 to 3 cm long at

one end.

ECOLOGY

Calotropis gigantea grows in dry uncultivated land, open waste land, along

road sides and railways, up to 1000 m altitude.

It grows on a variety of soils, but prefers sandy soils. Grow in different

climates, but usually with a periodic dry period.

HARVESTING

The leaves, flowers and roots of Calotropis gigantea are harvested

through out the year.

YIELD

Calotropis gigantea reaches a maximum height of 166 cm in 1 year,

producing 7.3 tons of fresh leaves and 56 kg latex per hectare.

P a g e | 30

SUBSTITUTES

In Asia Calotropis gigantea is used as a substitute for ipecacuanha

(Carapichea ipecacuanha (Brot.) L.Andersson), from tropical America, as an

effective cure for amoebic dysentery, but it has a stronger tendency to produce

vomiting and depression.

CHEMICAL CONSTITUENTS27

Aerial parts: Calotropin, calotoxin, uscharin,voruscharin, uschridin,

uzarigenin, syriogenin, calotonic acid and proceroside.

Flowers: Cyanidine-3-rhamnoglucoside

Leaves: Two new cardenolides (19-Nor- and 18, 20-epoxy-cardinolides)

Root bark: Three cardenolide glycosides, calotropin, frugoside, and

4'-O-beta-Dglucopyranosylfrugoside.

Giganticine, a non protein amino acid.

Roots: Two new oxypregnaneoligo glycosides

calotroposides A and B Clot inducing and dissolving activity

The latex of the plant C. gigantea hydrolyzed casein, fibrinogen and crude

fibrin in dose dependent manner. The proteolytic action on fibrinogen subunits was in

the order of Alpha > Beta > Gamma. The extracts also hydrolyzed all the subunits

(alpha polymer, alpha - chains, gamma - gamma dimmer and beta - chain) of

fibrin efficiently. The latex of C. gigantea exhibited procoagulant activity as

assayed by re-calcification time. In addition, the extracts dose- dependently

hydrolyzed blood and plasma clots. The proteolytic activity of C. gigantea latex

extract on different substrates was inhibited by IAA (Indole acetic acid).

P a g e | 31

PHARMACOLOGY27

ANTI-DIARRHOEAL ACTIVITY

The hydro alcoholic (50:50) extract of aerial parts of C. gigantea

showed significant anti-diarrhoeal activity against castor oil induced diarrhoea

model in rats. The ac- tivity was comparable to atropine (3mg/kg, i.p.). The plant

extract at a dose of 200 and 400 mg/kg i.p. showed significant reduction in fecal

output and frequency of defaecation when compared with castor oil treated rats.

The plant extracts also significantly inhibited castor oil induced enteropooling,

intestinal transit, weight and volume of intestinal content in all the doses .

Anti-diarrhoeal activity of ethanolic extract of C. gigantea flowers against castor

oil induced diarrhoea model in rats have also been reported and as per the study the

extract reduced the number, frequency and wetness of faeces as well as the propulsion

of charcoal meal .

ANTIOXIDANT ACTIVITY

The latex as well as the leaf extract of C. gigantea was investigated for its

antioxidant activity. The latex of C. gigantea (10mg/ml) exhibited greater

capacity to scavenge DPPH radicals, where as leaf extract showed moderate free

radical scavenging activity, which is proved to have anti- oxidant activity .

ANTI-FERTILITY ACTIVITY

The C. gigantea roots were examined for its Anti- fertility activity. The

ethanol extract and its chloroform- soluble fraction of C. gigantea roots exhibited

100% pregnancy interceptive activity in rats when administered as a single oral

dose of 100 mg/kg on day one post coitum. It was also found 7-25% loss in body

weight at the minimum effective contraceptive dose (MED) in rats treated with the

ethanolic extract as well as its chloroform soluble fraction.

P a g e | 32

ANTI-INFLAMMATORY ACTIVITY26

Anti- inflammatory activity of C. gigantea against carrageenan and kaolin

induced paw oedema in rats for acute - inflammatory condition, cotton-pellet

granuloma and adjuvant-arthritis model for chronic inflammatory condition has been

reported. The alkaloid fraction was found to possess comparatively high initial anti-

inflammatory activity. The residual anti- inflammatory activity of alkaloid fraction of

C. gigantea suggest either a greater protein binding nature of the compound there by

providing a slow released pool of active drug molecule in the system or non available

of possible bioactive metabolites to retain the activity pro- file relation .

ANALGESIC ACTIVITY

The ethanol extract of the flowers of C. gigantea was evaluated for the

assessment of its Analgesic activity using acetic acid induced writhing test model and

hot plate model in mice. The ethanol extract of the flowers of C. Gigantea showed

significant dose dependent analgesic activity against both the models i.e.

chemical (acetic acid induced writhing test) and thermal (hot plate method)

models in mice .

ANTI-PYRETIC ACTIVITY

The C. gigantea roots were studied for its antipyretic activity against yeast

induced pyrexia in rats and TAB (Typhoid) vaccine- induced pyrexia in rabbits. The

intraperitoneal administration of root extract at a dose of 200 and 400 mg/kg body

weight showed significant dose dependent anti-pyretic activity against both the

models.

ANTICANCER AND CYTOTOXIC ACTIVITY

Three cardenolide glycosides namely calotropin, frugoside, and 4'-O-beta-

D-glucopyranosyl frugoside were obtained as the cytotoxic principles from

"akondmul" (roots of Calotropis gigantea L.). They were found to be toxic to cell

lines of human origin, but not to those from mouse at 2 micrograms/ml .

P a g e | 33

ANTI CONVULSANT ACTIVITY

The alcoholic extract of the peeled roots of "akondmul" (Calotrophis

gigantea) showed a significant dose dependent anti-convulsant activity against

pentylenetetrazole (PTZ) induced convulsions in rats. Animals treated with the

extract showed anti anxiety activity and spent more time in the open arm of EPM. It

was also reported that the extract treated rats exhibited decreased locomotor

activity and fall-off time (motor coordination). The potentiation in the

pentobarbitone induced sleep due to the sedative effect of the extract was also

reported in the extract treated animals .

OTHER USES

The leaf extracts of Calotrophis gigantea showed significant inhibition

against mitochondrial malate dehydrogenase and malic enzyme of a filarial worm

Setaria digitata .The ethanol extract of Calotrophis gigantea showed antibacterial

activity against both gram positive and gram negative bacteria.

MEDICINAL USES

The whole plant is used for skin diseases, boils and sores and as a tonic and

purgative in small doses, and as an emetic in larger doses.

The powdered root bark is used to cure dysentery, elephantiasis, and leprosy.

The stem bark is diaphoretic, expectorant and used in the treatment of

dysentery, convulsions, lumbago, scabies, ringworm, pneumonia and to

induce labour.

The latex is used on stings, toothache, caries, ringworm, leprosy, syphilis,

rheumatism and tumours, and also as an antiseptic, vermifuge, emetic and

purgative.

The powdered flowers are given for cough, cold and asthma. The crushed

and warmed leaves are applied on burn, headache and rheumatic pain, and

as a tincture for intermittent fever.

Flower infusion is taken to treat intestinal worms, rheumatism and epileptic

attacks.

P a g e | 34

ACUTE DERMAL TOXICITY – FIXED DOSE PROCEDURE36

The acute dermal toxicity study was carried out in adult female albino rats by

“fix dose” method of OECD (Organization for Economic Co-operation and

Development) Guideline 434. Latex of the plant Calotropis gigantea was applied

topically at dose level 2000 mg/kg. No abnormal signs were observed at this dose

level.

FATAL DOSE24 Not determined

FATAL PERIOD24 Varies from half-an-hour to eight hours.

POSTMORTEM APPEARANCE

Signs of irritation in the stomach and intestine may be seen. Dilated pupils,

froth at the nostrils, stomatitis and inflammation of the gastro intestinal tract. The

abdominal viscera and brain are congested.

MEDICO LEGAL POINTS

The milky juice used as a vesicant, as a depilatory and as a remedy for chronic

skin affections Madar juice is occasionally used for the purpose of suicide,

infanticide and homicide.

P a g e | 35

ALLIUM CEPA (VENGAYAM)

TAXONOMICAL CLASSIFICATION

Kingdom : Plantae

Division : Angiosperms

Class : Monocots

Order : Asparagales

Family : Alliaceae

Genus : Allium

Species : cepa

VERNACULAR NAMES

English : Onion

Malayalam : Cyvannulli

Telugu : Erragadda

Hindi : Pyaj

Tamil : Venkayam, Ulligadda

Kannada : Niruli

Sanskrit : Polanduh

P a g e | 36

DESCRIPTION

A perennial herb, strong smelling when crushed; bulbs vary in size and shape

from cultivation to cultivation, often depressed globose and up to 20 cm in diameter;

outer tunics membranous. Stem up to 100cm tall and 30 mm in diameter, tapering

from inflated lower part. Leaves up to 40 cm in height and 20mm in diameter, usually

almost semicircular in section and slightly flattened on upper side basal in first year,

in second year their bases sheathing the lower sixth of the stem. Umbel 4 to 9cm in

diameter, subglobose or hemispherical, dense, many flowered pedicels up to 40mm,

almost equal. Perianth, stellate segments 3 to 4.5 × 2 to 2.5mm, white, with green

stripe, slightly unequal, the outer ovate, the inner oblong, obtuse or acute. Stamens

exerted; filaments 4 to 5mm, the outer subulate, the inner with an expanded base up to

2 mm wide and bearing short teeth on each side. Ovary is whitish Capsule.

GENERAL APPEARANCE

Macroscopically, Allium cepa varies in size and shape from cultivation to

cultivation, 2 to 20cm in diameter; flattened, spherical or pear shaped; white or

coloured.

GENERAL IDENTITY TESTS

Macroscopic inspection, microscopic characteristics and micro chemical

examination for organic sulfur compounds and thin- layer chromatographic analysis

for the presence of cysteine sulfoxides.

MICROSCOPIC CHARACTERISTICS

The external dried leaf scales of the bulbs show a large-celled epidermis with

lightly spotted cell walls; the cells are elongated longitudinally. The underlying

hypodermis runs perpendicular to the epidermis and contains large calcium oxalate

crystals bordering the cell walls. Large calcium oxalate crystals are found in the

hypodermis; stomata rare; large cell nuclei conspicuous; and spiral vessel elements

occur in the leaf mesophyll.

P a g e | 37

ORGANOLEPTIC PROPERTIES

Odour strong, characteristic alliaceous; taste strong; crushing or cutting the

bulb stimulates lacrimation.

POWDERED PLANT MATERIAL

Contains mainly thin walled cells of the mesophyll with broken pieces of

spiral vessel elements; cells containing calcium oxalate crystals are seen.

MAJOR CHEMICAL CONSTITUENTS

Sulfur and non-sulfur containing chemical constituents have been isolated

from Allium Cepa; the sulfur compounds are the most characteristic.

The organic sulfur compounds of Allium cepa, including the thiosulfinates,

cepaenes, S-oxides, S,S'-dioxides, monosulfides, disulfides, trisulfides, and

zwiebelanes occur only as degradation products of the naturally occurring cysteine

sulfoxides (e.g. (+)-S-propyl-L-cysteine sulfoxide). When the onion bulb is crushed,

minced, or otherwise processed, the cysteine sulfoxides are released from

compartments and contact the enzyme alliinase in adjacent vacuoles.

Hydrolysis and immediate condensation of the reactive intermediate

(sulfenic acids) are odorous thiosulphonates occur (in low concentrations) only in

freshly chopped onions, whereas the sulfides accumulate in stored extracts or

steamdistilled oils. Approximately 90% of the soluble organic-bound sulfur is present

as γ-glutamylcysteine peptides, which are not acted on by alliinase. They function as

storage reserve and contribute to the germination of seeds. However, on prolonged

storage or during germination, these peptides are acted on by γ-glutamyl

transpeptidase to form alk(en)yl-cysteine sulfoxides, which in turn give rise to other

volatile sulfur compounds.

Among the flavonoids, onions also provide a particularly large amount of

quercetin. A wide variety of allyl sulfides are found in onion, including the four major

diallyl sulfides: DMS (diallyl monosulfide), DDS (diallyl disulfide), DTS (diallyl

trisulfide), and DTTS (diallyl tetrasulfide). Also present are a wide variety of

P a g e | 38

sulfoxides, including S-methyl-L-cysteine sulfoxide (MCSO), S-(1-propenyl)-L-

cysteine sulfoxide (PRENCSO), S-methyl- l-cysteine sulfoxide, S-propyl- l-cysteine

sulfoxide, and S-propenyl- l-cysteine sulfoxide.

Onions are a very good source of immune-supportive vitamin C. They are

also a good source of enzyme-activating manganese and molybdenum as well as

heart-healthy vitamin B6, fiber, folate, and potassium.

RAW ONION

Nutritional value per : 100 g (3.5 oz)

Energy : 166 kJ (40 kcal)

Carbohydrates : 9.34 g

Sugars : 4.24 g

Dietary fibre : 0.7 g

Fat : 0.1 g

Saturated fat : 0.042 g

Monounsaturated fat : 0.013 g

Polyunsaturated fat : 0.017 g

Protein : 1.1 g

Water : 89.11 g

Vitamin A : 0.04g (0%)

Thiamine (Vit.B1) : 0.046 mg (4%)

Riboflavin (Vit.B2) : 0.027 mg (2%)

Niacin (Vit.B3) : 0.116 mg (1%)

Vitamin B6 : 0.12 mg (9%)

Folate (Vit.B9) : 19g (5%)

Vitamin B : 120g (0%)

Vitamin C : 7.4 mg (12%)

Vitamin E : 0.02 mg (0%)

Vitamin K : 0.4g (0%)

Calcium : 23 mg (2%)

Iron : 0.21 mg (2%)

Magnesium : 0.129 mg (0%)

Phosphorus : 29 mg (4%)

P a g e | 39

STORAGE FORMS

Dried Allium cepa products should be stored in well closed containers,

protected from light, moisture, and elevated temperature. Fresh bulbs and juice should

be refrigerated (2 to 10°C).

PHARMACOLOGICAL ACTIVITIES30

ANTI-INFLAMMATORY

In onion a unique sulfur molecule is found in the bulb portion of the plant has

been shown to inhibit the activity of macrophages.

Onion's antioxidants--including flavonoid antioxidant, quercetin have anti-

inflammatory benefits. These antioxidants help to prevent the oxidation of fatty acids

in our body. When we have lower levels of oxidized fatty acids, our body produces

fewer pro- inflammatory messaging molecules, and our level of inflammation is kept

in check.

The organosulphur compounds in these spices inhibit the oxidation of fatty

acids, thereby preventing the formation of pro- inflammatory messengers, and inhibit

bacterial growth, via interaction with sulphur-containing enzymes.

Topical application of an aqueous extract of Allium cepa (10% in a gel

preparation) inhibited ear oedema induced by arachidonic acid in mice. The active

anti-allergic and anti- inflammatory constituents of onion are the flavonoids (quercetin

and kaempferol).The flavonoids act as anti- inflammatory agents because they inhibit

the action of protein kinase, phospholipase A2, cyclooxygenase, and lipoxygenase , as

well as the release of mediators of inflammation (e.g. histamine) from leukocytes.

P a g e | 40

CLINICAL STUDIES30

EFFECTS ON SKIN

The effectiveness of topical use of crude onion juice in the treatment of

patchy alopecia areata in comparison with tap water was tested. At four weeks, hair

re-growth was seen in 17 patients (73.9%), and at six weeks, the hair regrowth was

observed in 20 patients (86.9%) and was significantly higher among males (93.7%)

compared to females (71.4%) p<0.0001. In the tap-water treated-control group, hair

regrowth was apparent in only 2 patients (13%) at 8 weeks of treatment with no sex

difference.

ANTI - CARCINOGENIC AND ANTI - MUTAGENIC ACTIVITIES

Onion consumption is associated with reduced risk of developing brain cancer,

as confirmed in a hospital-based case-control study conducted in the North East

China, on a total of 129 histologically confirmed brain cancer cases.

In the Netherlands Cohort Study, comprising 1, 20,852 men and women aged

between 55 and 69 reported a strong association between onion consumption and a

reduction in the incidence of stomach carcinoma. However, in the same study group,

the authors found no evidence for a protective effect of Allium vegetable consumption

and reduced risk of developing colon, rectum carcinoma, lung or female breast

cancer.

CARDIOVASCULAR EFFECTS

In an open, randomized placebo-controlled, double-blind, cross-over phase I

study in 10 apparently healthy volunteers (8 men and 2 women), a decrease in arterial

blood pressure, a reduction in plasma viscosity and haematocrit were observed 5

hours after administration of an onion- olive - oil macerate (corresponding to a mean

daily dose of 2.5 g fresh onion). There was a tendency to reduce by 5 to 8% in systolic

and by 5 to 10% in diastolic blood pressure. There was still a minimal reduction of

blood pressure detectable 8 hours after administration. The mean reduction of 1.6% in

haematocrit and 0.04 mPa/s in plasma viscosity were measured.

P a g e | 41

ANTI - AGGREGATORY ACTIVITY

Onions, fried or boiled, not only prevented reduction, but also caused a

marked increase of fibrinolytic activity. Decalcified clotting times, thromboses, and

cholesterol and fibrinogen levels were not significantly changed.

The effect of onion juice consumption on some blood properties, influenced

by consumption of 100 g butter, has been studied in 10 healthy adult males, between

35 and 50 years old. The juice was prepared freshly from 50 g of onion and it was

administered orally as a single dose. After administration of onion juice, the blood

coagulation time (which had decreased by 13.3% after ingestion of butter) was

increased by 9.8%. Analogically, fibrinolytic activity (decreased by 48.6%) increased

by 15.7%. Examined the effects of onion consumption (70 g of raw onion daily for a

period of 7 days) on platelet thromboxane (TXB2) production in five women in the

age range of 25 to 65 years. Onion feeding slightly but not significantly increased

amount of TXB2, from mean 910 to 1005 pmol/ml of serum.

ANTI - DIABETIC ACTIVITY

A comparative study, using a cross over design, was carried out on 20 well

controlled diabetic out - patients to investigate the metabolic effects of the diet (68%

calorie carbohydrate, 20% calorie fat, 12% calorie protein) and 3 x 20 g fresh o nion

per day during two weeks. Study resulted in significant decrease in blood sugar level

(4.37 mg%, p<0.05).

LIPID - LOWERING EFFECTS

The effect of onion on alimentary hyperlipidemia, induced by feeding 100 g

butter, has been studied in 10 healthy adult males, between age of 35 and 50 years.

The juice was prepared freshly from 50 g of onion and was administered orally as a

single dose. Administration of onion juice completely prevented the rise in serum

cholesterol in fact slightly lowered it.

P a g e | 42

USES

Allium cepa is used in treatment of cold, allergies, toothaches, laryngitis and

coughing. Healers make a tincture of onion to treat a variety of conditions

including diarrhea, facial paralysis, hay fever, hernia, laryngitis, pneumonia and

trauma . Onion has been used for healing both internally and externally. Internally,

onion has been recommended to treat bronchitis, whooping cough, asthma and

other respiratory problems. It has believed to help loosen congestion in the lungs

and expand the airways.

TRIBAL CLAIMS

A mixture of rue (Ruta graveolens) and onion is used to rid the digestive

system of parasites. Onion is also thought to stimulate the appetite. It is also thought

to help reduce arteriosclerosis by lowering blood cholesterol levels and preventing the

formation of blood clots. Externally, fresh onion juice is used to prevent bacterial and

fungal infections. It can be applied to wounds and stings on the skin, used to

remove warts and even to reduce unwanted skin blemishes. Warm onion juice

dropped in the ear is said to help relieve earache.

ACUTE TOXICITY STUDY31

Swiss albino mice of either sex weighing (18 to 22g) and of 90 days age were

used for acute oral toxicity study. The study was carried out as per the guidelines set

by OECD. The animals were starved overnight and divided into six groups (n=3)

and were fed with increasing doses (10, 30, 100, 300, 1000, 2000, 3000 mg/kg B.W.)

of the petroleum ether, solvent ether, chloroform, alcohol and chloroform water

extract. The animals were continuously observed for mortality and behavioral

responses for 48 h and thereafter once daily for 14 days after administration. Upto

3000 mg no mortality was observed.

P a g e | 43

ADVERSE REACTIONS

Allergic reactions such as rhino-conjunctivitis and contact dermatitis have

been reported.

PRECAUTIONS

No general precautions have been reported, and no precautions have been

reported concerning drug interactions, drug and laboratory test interactions, nursing

mothers, pediatric use.

P a g e | 44

CHAPTER IV

4.1. COLLECTION, AUTHENTICATION, PURIFICATION

AND PREPARTION OF “POORA PARPAM”

COLLECTION OF DRUG:

The raw drug was collected from country drug merchant shop,

Chennai

AUTHENTICATION:

The raw drug Pooram was authenticated by Siddha Central

Research Institute, Chennai.

PURIFICATION OF POORAM1

Pooram (raw) – 35 gm,

Vettrilai (Piper bettle) leaves – 8.75gm,

Milagu (Piper nigram) – 8.75gm,

METHOD OF PURIFICATION:

Piper bettle leaves and Piper nigram seeds were ground together

and made into a poultice. Then one liter of water was taken in a mud pot and

the poultice was mixed in that water. Pooram (raw) was covered with a piece

of clean dry cloth, so that it was not exposed outside. One end of the cloth was

tied to a bamboo stick and placed horizontally over the opening of the mud

pot. The raw drug Pooram in cloth was suspended in the above decoction. The

vessel was constantly heated till decoction reduced by three fourth of its

volume. Finally the Pooram was taken out from the cloth, washed with clean

water and dried in sunlight.

- Gunapadam Thathu Jeeva Vaagupu, 283, 284

P a g e | 45

PREPARATION OF POORA PARPAM

INGREDIENTS:

Purified Pooram (Calomel) – ¾ varaagan (13.65 gm),

Erukkam paal

(Latex juice of Calotrophis gigentia) – 14 palam (490gm),

Vengayam (Juice of Allium cepa) – 14 palam (910ml)

METHOD OF PREPARATION:

Purified pooram was placed in a clean dry cloth, fully covered and

tied together. In a mud pot the above mentioned quantity of Erukkam paal was

taken and Pooram covered by cloth was placed inside the mud pot. The mud

pot was constantly heated until latex juice remaining in the pot totally dried

out. Pooram was removed from the cloth and washed with water. Then it was

ground into paste with Vengayam juice and made into a poultice. Small pills

were made from the poultice and dried in shade. Pills were placed inside a

mud plate and enclosed with similar size plate. Margins of the plate were

covered with clay pasted cloth. Finally it was completely dried, kept in a pit

under earth and puddam was made with 40 palam cow dung cakes (1400 gms).

Next morning on being cooled mud plate was removed and finished Poora

Parpam was separated and ground into a fine powder. Later Parpam was

collected in a clean air tight container.

Dose of drug : One Kadugu (mustard seed)

Adjuvant : Butter / Ghee.

Therapeutic uses : Kirandhi noi, Natpatta Soolai, Natpatta Keelvayu

Contra indications : Avoid salt and tamarind.

- Veerama Munivar Vagada Thiratu - 69,70

P a g e | 46

Fig.1. UNPURIFIED POORAM

Fig.2. PURIFIED POORAM

P a g e | 47

Fig.3. POORA PARPAM

P a g e | 48

4.2. QUALITATIVE ANALYSIS

PHYSICO-CHEMICAL PROPERTIES

COLOUR

About 50 gm of Poora Parpam was taken in a clean glass beaker

and tested for its colour by viewing against a white opaque back ground

under direct sunlight.

ODOUR

About 50 gm of the Poora Parpam was placed in 100 ml of

beaker and tested for its odour by wafting the air above the beaker.

pH

The ph of the Poora Parpam was estimated as per the method

prescribed in the Indian standard (IS) - 6940(1982). One gram of the

Poora Parpam was taken into a 100ml graduated cylinder containing

about 50 ml of water and filled up to the mark with water. The cylinder

was stopped and shaken vigorously for two minutes and the suspension

was allowed to settle for an hour at 25 to 27˚c. About 25 ml of the clear

aqueous solution was transferred into a 50 ml beaker and tested for pH

using DIGISUN digital PH meter (DIGISUN electronics, Hyderabad,

India)

DETERMINATION OF ASH VALUE

Two gms of the Poora Parpam was weighed accurately in tarred

platinum or silica dish and incinerated at a temperature not exceeding

450˚c until free from carbon, then cooled and weighed. Calculate the

percentage of ash with reference of the air dried drug.

P a g e | 49

WATER SOLUBLE ASH

To the Gooch crucible containing the total ash, 25 ml of water was

added and boiled for 5 minutes. The insoluble matter was collected in a

sintered glass crucible or on ash less filter paper. Washed with hot water

and ignited in a crucible for 15 minutes at a temperature not exceeding

450˚c. Subtract the weight of the insoluble matter from the weight of the

ash. The difference of the weight represents the water soluble ash.

Calculate the percentage of water soluble ash with reference to the air

dried drug.

ACID INSOLUBLE ASH

Ash was boiled for 5 minutes with 25 ml of 1:1 diluted. Hcl. The

insoluble matter was collected in a Gooch crucible and placed on an ash

less filter paper, washed with water and then ignited. Finally cooled in a

desiccator and weighed. The percentage of insoluble ash was calculated

with reference to the air dried drug.

LOSS ON DRYING

Five grams of the Poora Parpam was heated in a hot oven at 105˚c

to a constant weight. The percentage loss of weight was calculated as per

procedure.

P a g e | 50

Table .1. CHEMICAL ANALYSIS

EXPERIMRNT

OBSERVATION

INFERENCE

UNPURIFIED

POORAM

PURIFIED

POORAM

Appearance of the sample

White in colour

White in colour

White in colour

Solubility:

a. A little of the sample is

shaken well with distilled

water

b. A little of the sample is

shaken well with

Con.HCl and Con.H2SO4

Insoluble

Insoluble

Absence of

silicate

Absence of

silicate

Action of heat:

A small amount of the

sample is taken in a dry

test tube and heated

gently at first and then

strongly

No colour fumes

Absence of

carbonate and

nitrate

Absence of

carbonate and

nitrate

Flame test:

The sample is mixed

with Con.HCL in a watch

glass and introduced in

luminous part of the

Bunsen flame

No colour flames

appeared

Absence of

copper

Absence of

copper

Ash Test:

A filter paper is soaked

into a mixture of sample

and cobalt nitrate solution

and introduced into the

Bunsen flame and ignited

No yellow colour

flame

Absence of

sodium.

Absence of

sodium.

P a g e | 51

PREPARATION OF EXTRACT:

5g of sample was taken in a 250 ml of clean beaker and 50 ml of distilled

water was added to it. Then it was boiled well for about 10 min. Then it is allowed to

cool and filtered in a 100 ml volumetric flask and made up to 100 ml with distilled

water. This preparation is used for the qualitative analysis of acidic and basic radicals.

TEST FOR BASIC RADICALS

PROCEDURE

OBSERVATION

UNPURIFIED

POORAM

PURIFIED

POORAM

Test for Potassium:

A pinch of sample is

treated with 2ml of

sodium nitrate solution

and then treated with 2ml

of cobalt nitrate in 30%

of glacial acetic acid.

No formation of

yellow colour

precipitate

Absence

of Potassium

Absence of

Potassium

Test for Calcium:

2 ml of extract is taken in

a clean test tube. To this

add 2 ml of 4%

ammonium oxide

solution.

No formation of

white colour

precipitate

Absence of

Calcium

Absence of

Calcium

Test For Magnesium:

To 2ml of extract,

Sodium hydroxide

solution is added in

drops.

Formation of white

colour precipitate

Presence of

Magnesium

Presence of

Magnesium

P a g e | 52

Test For Ammonium:

To 2ml of extract few ml

of Nessler's reagent and

excess of sodium

hydroxide solution are

added.

No appearance of

Brown colour

Absence of

Ammonium

Absence of

Ammonium

Test For Sodium:

2 pinches of the sample is

made into paste by using

Hcl and introduced into

the blue flame of Bunsen

burner.

No Characteristic

changes

Absence of

Sodium

Absence of

Sodium

Test for Iron (Ferrous):

The extract is treated with

Conc. HNO3 and

ammonium thiocynate.

Appearance of

Blood red colour

Presence of

Ferrous iron

Presence of

Ferrous iron

Test For Zinc:

To 2m1 of the extract

sodium hydroxide

solution is added in

drops.

No Formation of

White colour

precipitate

Absence of

Zinc

Absence of

Zinc

Test For Aluminium:

To the 2m1of the extract

sodium hydroxide is

added in drops.

No Characteristic

changes

Absence of

Aluminium

Absence of

Aluminium

P a g e | 53

Test For Lead:

2 ml of extract is added

with 2ml of potassium

iodide solution.

Formation of yellow

colour precipitate

Absence of

Lead

Absence of

Lead

Test for Copper:

a. One pinch of substance

is made into paste with

con. Hcl in a watch glass

and introduced into the

non- luminous part of the

flame.

b. 2 ml of extract is

added with excess of

ammonia solution.

No Formation of

Blue colour

Precipitate.

No Formation of

Blue colour

Precipitate

Absence

of Copper

Absence

of Copper

Test For Mercury:

2m1 of the extract is

treated with 2ml of

sodium hydroxide

solution.

Formation of

Yellow precipitate

Presence

of Mercury

Presence

of Mercury

Test for Arsenic:

2m1 of the extract is

treated with 2ml of

sodium hydroxide

solution.

No Formation of

Brownish red

precipitate

Absence

of Arsenic

Absence

of Arsenic

P a g e | 54

TEST FOR ACID RADICALS

PROCEDURE

OBSERVATION

UNPURIFIED

POORAM

PURIFIED

POORAM

Test for Sulphate:

2 ml of the extract is

added to 5 % barium

chloride solution.

No formation of

white precipitate

Absence

of Sulphate

Absence

of Sulphate

Test for Chloride:

The extract is treated

with Silver nitrate

solution.

No Formation of

White precipitate

Absence

of Chloride

Absence

of Chloride

Test for Phosphate :

The extract is treated

with ammonium

molybdate and conc.

HNO3.

No formation of

Yellow precipitate

Absence

of Phosphate

Absence

of Phosphate

Test for Carbonate :

The substance is

treated with Conc.

HCl.

No Formation of

Effervescence

Absence of

Carbonate

Absence of

Carbonate

Test for fluoride &

oxalate:

2ml of extract is added

with 2ml of dil.acetic

acid and 2ml calcium

chloride solution and

heated.

No Formation of

cloudy appearance

Absence of

Fluoride &

Oxalate

Absence of

Fluoride &

Oxalate

P a g e | 55

OTHER CONSTITUENTS

PROCEDURE

OBSERVATION

INFERENCE

UNPURIFIED

POORAM

PURIFIED

POORAM

Test for Starch:

The extract is added

with weak iodine

solution.

Formation of blue

colour

Presence of

Starch

Presence of Starch

Test for Reducing

Sugar:

5 ml of Benedict's

qualitative solution

is taken in a test

tube and allowed to

boil for 2 min.

Add 8 to 10 drops

of extract and again

boil it for 2min. The

colour changes are

noted.

No Colour changes

Absence of

Reducing

sugar

Absence of Reducing

sugar

Test For Nitrate:

1gm of the substance

is heated with copper

turnings and

concentrated H2SO4

and viewed the test

tube vertically down.

No Characteristic

changes

Absence of

Nitrate

Absence of

Nitrate

P a g e | 56

Test for

Alkaloids:

a. 2ml of the extract

is treated with 2ml

of potassium Iodide

solution

b. 2m1 of extract is

treated with 2ml of

picric acid

c. 2m1 of the

extract is treated

with 2ml of

phosphotungstic

acid

No Appearance of

Red colour

No Appearance of

Yellow colour

No Appearance of

white precipitate

Absence of

Alkaloids

Absence of

Alkaloids

Absence of

Alkaloids

Absence of

Alkaloids

Absence of

Alkaloids

Absence of

Alkaloids

Test for amino

acids :

Dilute extract + 2ml

of Ninhydrin’s

solution.

No Appearance of

violet colour

Absence of

Amino acids

Absence of Amino

acids

Test for Tannic

acid :

The extract is

treated with Ferric

chloride.

No formation of

Blue black

precipitate

Absence of

Tannic acid

Absence of Tannic

acid

P a g e | 57

Test for type of

Compound:

2ml of the extract is

treated with 2 ml of

ferric chloride

solution

No Appearance of

Green colour

No Appearance of

Red colour

No Appearance of

Violet colour

No Appearance of

Blue colour

Absence of

Oxyquinole,

Epinephrine and

Pyro catechol

Absence of Anti

-pyrine,

Aliphatic amino

acids and

Meconic acid

Absence of

Apomorphine

Salicylate and

Resorcinol

Absence of

Morphine,

Phenol

cresol and

Hydroquinone

Absence of

Oxyquinole,

Epinephrine and

Pyro catechol

Absence of Anti

pyrine, Aliphatic

amino acids,

Meconic acid

Absence of

Apomorphine

Salicylate and

Resorcinol

Absence of

Morphine, Phenol

Cresol,

Hydroquinone

P a g e | 58

TABLE.2. CHEMICAL ANALYSIS

EXPERIMRNT

OBSERVATION

INFERENCE

POORAM

PARPAM

Appearance of the sample

White in colour

White in colour

Solubility:

a. A little of the sample is

shaken well with distilled water

b. A little of the sample is

shaken well with con.HCl

con.H2SO4

Insoluble

Absence of silicate

Action of heat:

A small amount of the sample is

taken in a dry test tube and

heated gently at first and the

strongly

No colour fumes

Absence of

carbonate and

nitrate

Flame test:

The sample is mixed with

Con.HCL in a watch glass and

introduced in luminous part of

the Bunsen flame

No colour flames appeared

Absence of copper

Ash Test:

A filter paper is soaked into a

mixture of sample and cobalt

nitrate solution and introduced

into the Bunsen flame and

ignited

No yellow colour flame

Absence of sodium.

P a g e | 59

Preparation of extract:

5g of sample was taken in a 250 ml of clean beaker and 50 ml of distilled

water was added to it. Then it was boiled well for about 10 min. Then it is allowed to

cool and filtered in a 100 ml volumetric flask and made up to 100 ml with distilled

water. This preparation is used for the qualitative analysis of acidic and basic radicals.

TEST FOR BASIC RADICALS

PROCEDURE

OBSERVATION

INFERENCE

POORAM

PARPAM

Test for Potassium:

A pinch of sample is treated

with 2ml of sodium nitrate

solution and then treated with

2ml of cobalt nitrate in 30% of

glacial acetic acid.

No formation of Yellow

colour precipitate

Absence of

Potassium

Test for Calcium:

2 ml of extract is taken in a

clean test tube. To this add 2 ml

of 4% ammonium oxide

solution.

No formation of white

colour precipitate

Absence of

Calcium

Test For Magnesium:

To 2ml of extract, Sodium

hydroxide solution is added in

drops.

Formation of White

colour precipitate

Presence of

Magnesium

P a g e | 60

`Test For Ammonium:

To 2ml of extract few ml of

Nessler's reagent and excess of

sodium hydroxide solution are

added.

No appearance of Brown

colour

Absence of

Ammonium

Test For Sodium:

2 pinches of the substance is

made into paste by using Hcl

and introduced into the blue

flame of Bunsen burner.

No Characteristic

changes

Absence of

Sodium

Test for Iron (Ferrous) :

The extract is treated with Conc.

HNO3 and ammonium

thiocynate.

Appearance of Blood red

colour

Presence of

Ferrous iron

Test For Zinc:

To 2m1 of the extract sodium

hydroxide solution is added

indrops.

No Formation of White

colour precipitate

Absence of

Zinc

Test For Aluminium:

To the 2m1of the extract sodium

hydroxide is added in drops.

No Characteristic

changes

Absence of

Aluminum

Test For Lead:

2 ml of extract is added with

2ml of potassium iodide

solution.

Formation of yellow colour

precipitate

Absence of

Lead

Test for Copper:

a. one pinch of substance is

made into paste with con. Hcl in

a watch glass and introduced

No Formation of Blue

colour Precipitate.

Absence

of Copper

P a g e | 61

into the non- luminous part of the

flame.

b. 2 ml of extract is added with

excess of ammonia solution.

Test For Mercury:

2m1 of the extract is treated

with 2ml of sodium hydroxide

solution.

Formation of Yellow

precipitate

Presence

of Mercury

Test for Arsenic:

2m1 of the extract is treated

with 2ml of sodium hydroxide

solution.

No Formation of Brownish

red precipitate

Absence

of Arsenic

TEST FOR ACIDIC RADICALS

PROCEDURE

OBSERVATION

POORAM

PARPAM

Test for Sulphate :

2 ml of the extract is added to 5

% barium chloride solution.

No formation of white

precipitate

Absence of

Sulphate

Test for Chloride :

The extract is treated with Silver

nitrate solution.

No Formation of White

precipitate

Absence of

Chloride

P a g e | 62

Test for Phosphate :

The extract is treated with

ammonium molybdate and conc.

HNO3.

Formation of

Yellow precipitate

Presence of

Phosphate

Test for Carbonate :

The substance is treated with

Conc. HCl.

No Formation of

Effervescence

Absence of

Carbonate

Test for fluoride & oxalate:

2ml of extract is added with 2ml

of dil.acetic acid and 2ml

calcium chloride solution and

heated.

No Formation of cloudy

appearance

Absence of

Fluoride & Oxalate

Test For Nitrate:

1gm of the substance is heated

with copper turnings and

concentrated H2SO4 and

viewed the test tube vertically

down

No Characteristic changes

Absence of Nitrate

P a g e | 63

OTHER CONSTITUENTS

PROCEDURE

OBSERVATION

INFERENCE

Test for Starch :

The extract is added with weak

iodine solution.

Formation of blue colour

Presence of Starch

Test for Reducing Sugar:

5 ml of Benedict's qualitative

solution is taken in a test tube

and allowed to boil for 2 min.

Add 8 to 10 drops of extract and

again boil it for 2min. The

colour changes are noted.

No Colour changes

Absence of

Reducing

sugar

Test for Alkaloids:

a. 2ml of the extract is treated

with 2ml of potassium Iodide

solution

b. 2m1 of extract is treated with

2ml of picric acid

c. 2m1 of the extract is treated

with 2ml of phosphotungstic

acid

No Appearance of

Red colour

No Appearance of Yellow

colour

No Appearance of

white precipitate

Absence of

Alkaloids

Test for amino acids :

Dilute extract + 2ml of

Ninhydrin’s solution.

No Appearance of violet

colour

Absence of Amino

acids

P a g e | 64

Test for Tannic acid :

The extract is treated with Ferric

chloride.

Formation of Blue black

precipitate

Presence of

Tannic acid

Test for type of Compound:

2ml of the extract is treated with

2 ml of ferric chloride solution

No Appearance of Green

colour

No Appearance of Red

colour

No Appearance of Violet

colour

No Appearance of Blue

colour

Absence of

Oxyquinole,

Epinephrine and

Pyrocatechol

Absence of Anti -

pyrine, Aliphatic

amino acids and

Meconic acid

Absence of

Apomorphine

Salicylate and

Resorcinol

Absence of

Morphine, Phenol

cresol and

Hydroquinone.

P a g e | 65

4.3. QUANTITATIVE ANALYSIS

EXPERIMENTAL PROCEDURE: DONE AT SAIF, IIT MADRAS.

HR SEM-METHODOLOGY:

An SEM is essentially a high magnification microscope, which uses a

focused scanned electron beam to produce images of the sample, both top -

down and, with the necessary sample preparation, cross-sections. The primary

electron beam interacts with the sample in a number of key ways:-

Primary electrons generate low energy secondary electrons, which tend to

emphasize the topographic nature of the specimen.

Primary electrons can be backscattered which produces images with a high

degree of atomic number (Z) contrast.

Ionized atoms can relax by electron shell- to-shell transitions, which lead to

either X-ray emission or Auger electron ejection. The X-rays emitted are

characteristic of the elements in the top few μm of the sample.

SAMPLE PREPARATION:

Sample preparation can be minimal or elaborate for SEM analysis,

depending on the nature of the samples and the data required. Minimal

preparation includes acquisition of a sample that will fit into the SEM chamber

and some accommodation to prevent charge build-up on electrically insulating

samples. Most electrically insulating samples are coated with a thin layer of

conducting material, commonly carbon, gold, or some other metal or alloy.

The choice of material for conductive coatings depends on the data to be

acquired: carbon is most desirable if elemental analysis is a priority, while

metal coatings are most effective for high resolution electron imaging

applications. Alternatively, an electrically insulating sample can be examined

without a conductive coating in an instrument capable of "low vacuum"

operation.

The SEM is carried out by using FEI-Quanta FEG 200-High

Resolution Instrument.

P a g e | 66

Resolution: 1.2 nm gold particle separation on a carbon substrate

Magnification: From a min of 12 X to greater than 1, 00,000 X

Application: To evaluate grain size, particle size distributions,

material homogeneity and inter metallic distributions.

INDUCTIVELY COUPLED PLASMA OPTICAL EMISSION

SPECTROMETRY (ICP-OES)

INTRODUCTION

Inductively coupled plasma optical emission spectrometry (ICP-OES),

is an analytical technique used for the detection of trace metals. It is a type of

emission spectroscopy that uses the inductively coupled plasma to produce

excited atoms and ions that emit electromagnetic radiation at wavelengths

characteristic of a particular element. The intensity of this emission is

indicative of the concentration of the element within the sample.

PRINCIPLE

A Perkin-Elmer Optima ICP spectrometer is used for routine ICP-OES

analysis. First, a high-energy radio frequency field is impinged upon a stream

of argon gas. Then, a spark is used to ionize the argon gas, which forms

sustained plasma due to inductive coupling with the high energy radio

frequency field and the continuous supply of fresh argon to the plasma torch.

This plasma has solutions passed into it in the form of a fine aerosol. The

aerosol is dried, the dried particles broken apart, and the individual elements

are excited by interaction with the excited state argon in the plasma. As each

atom returns to its ground state from the excited state, they emit light at

wavelengths characteristic of the elements from which they originate. The

emission intensity for each element is monitored for each standard solution

and a calibration curve of emission intensity versus element concentration ca n

be constructed.

P a g e | 67

EXTRACTION OF INFORMATION

Obtaining qualitative information, i.e., what elements are present in the

sample, involves identifying the presence of emission at the wavelengths

characteristic of the elements of interest. Obtaining quantitative information,

i.e., how much of an element is in the sample, can be accomplished using plots

of emission intensity versus concentration called calibration curves. Typical

calibration graph is illustrated below

Typical ICP Calibration curve

Perkin Elmer Optima 5300DV

40 M Hz RF generator;

Range: 165-782 nm;

Detection limit: Upto ppb level using SCD detector

P a g e | 68

SAMPLE PREPARATION – MICROWAVE DIGESTION

Weigh 0.25g of test sample and transfer into a liner provided with the

instrument.

Slowly add 9ml of Nitric acid or Sulphuric acid such that no piece of sample

sticks on the slides.

Mix thoroughly and allow reacting for few minutes.

Place the liner in the vessel jacket.

Close the screw cap hand-tight in clockwise direction.

Seal the vessel and place in the rotor fixed in microwave.

Set temperature to 180°C for 5 minutes; hold at 180°C for least 10 minutes.

Allow the vessels to cool down to a vessel interior temperature below 60°C

and to a vessel surface temperature (IR) below 50°C before removing the

rotor.

The digested sample was made up to 100ml with millipore water.

If visible insoluble particles exist, solution could be filtered through

Whatmann filter paper.

Transfer the digested solution into plastic containers and label them properly.

P a g e | 69

4.4. TOXICOLOGICAL EVALUVATION OF POORA PARPAM

INTRODUCTION:

Safety is a fundamental principle in the provision of traditional medicines and

herbal products for health care and a critical component of quality control. WHO

guidelines provide practical and technical guidance for monitoring the safety of

traditional medicines within pharmacovigilance systems. The safety monitoring of

traditional medicines is compared and contrasted with that of other medicines,

currently undertaken in the context of the WHO International Drug Monitoring

Programme. While there are regulatory and cultural differences in the preparation and

use of different types of medicines, they are all equally important from a

pharmacovigilance perspective.

SCOPE OF TOXICITY STUDY:

Siddha medicines are becoming increasingly popular as an effective and

relatively alternative to allopathic drugs. In order to make global acceptance the

quality and its safety are under scrutiny though traditional practitioners prescribed

them since long time.

In the recent past, several western research groups have high lightened these

pitfalls reporting the prevalence and concentration of heavy metals in Siddha

medicines .In Siddha system of medicine the formulations prepared with minerals are

called as herbo-mineral preparations such as Parpam, Chenduram etc, in traditional

language. It is mandatory to test raw materials used in Siddha formulations for the

presence of various toxic materials including heavy metal content.

All mineral preparations in Siddha, is prepared under special physico -

chemical processes that, according to the ancient Indian belief, 'detoxify', toxic heavy

metals in it. Strictly speaking, these constituents are thus not contaminants but

ingredients deliberately included for a specific curative purpose. India being a

signatory to WTO, to promote its products in the international market, it is imperative

to study their safety for human consumption.

P a g e | 70

PLAN OF WORK

The following studies were carried out on Poora Parpam

Acute toxicity

Long term toxicity

The toxicity studies were evaluated after getting permission from The

Institutional Animal Ethical Committee clearance. (1248/ac/09/CPCS EA/04/IAEC 2011).

ACUTE TOXICITY STUDY OF POORA PARPAM

PRINCIPLE

Acute toxicity was carried out in Swiss albino mice of either sex with a single

exposure of 10 times more than the recommended therapeutic dose of test drug and

the study duration was 14 days. The study was carried out as per the WHO guidelines.

Drug profile:

Test drug : Poora Parpam

Therapeutic dose : 12 mg/day.

Experimental animal:

Animal species : Swiss albino mice

Age / Weight : 6 weeks. Mice (20-25 gms).

Gender : Both male and female

Number of Animals : 20 (10-Male, 10- Female).

S.No Groups No of mice

1. Vehicle control 10 (5 male, 5 female)

2.

Acute dose 10 x therapeutic dose

(0.216 mg/animal) ( test group)

10 (5 male, 5 female)

P a g e | 71

TEST ANIMALS

Test animals were obtained from The King Institute, Chennai and kept at

animal house, National Institute of Siddha, Chennai. All the animals were kept under

standard environmental condition (22 ± 3˚c).The animals had free access to water and

standard pellet diet (Sai Meera foods pvt.ltd, Bangalore).The principles of laboratory

animal care were followed.

RANDOMIZATION, NUMBERING AND GROUPING OF ANIMAL

The animals were randomly divided into two groups. Each group consist of 10

animals (5 per sex in each group) the first groups kept as control and another group

was treated with test drug. The animals were allowed acclimatization period of 7 days

to laboratory conditions prior to the initiation of treatment. The females were

nulliparous and non pregnant.

IDENTIFICATION OF ANIMAL

By cage number, animal number and individual marking on fur with picric acid.

HOUSING & ENVIRONMENT

The animals were housed in polypropylene cages provided with bedding of

husk. Dark and light cycle each of 12 hours was maintained.

ROUTE OF ADMINISTRATION:

Oral route was selected, because it is the normal route of clinical

administration.

TEST SUBSTANCE AND VEHICLE

The Poora Parpam is white coloured, without taste and odour. The test

substance is insoluble in water, in order to obtain and ensure the uniformity in drug

distribution; the drug is dissolved by 10% aqueous tween 80 solution.

P a g e | 72

ADMINISTRATION OF DOSES

Poora Parpam was suspended in 10% aqueous tween 80 solution with uniform

mixing and it was administered to the test group in a single oral dose. The control

groups were received equal volume of the vehicle. The animals were weighed before

giving the drug. The dose level was calculated according to body weight and surface

area. Since the clinical dose was 12 mg/day it was converted to animal dose (0.216

mg/animal) and then administered to the test group animals.

OBSERVATIONS

Observations were made and recorded systematically and continuously as per

the WHO guideline after test drug administration. Animals were observed

individually for the first 4 hrs, then periodically during the study period. Mortality and

abnormal signs were observed during the study period. At the end of the study all the

animals were sacrificed and necropsy was done.

BODY WEIGHT

Individual weight of animals was determined before the test drug

administration and daily for 14 days.

.

LONG TERM TOXICITY STUDY OF POORA PARPAM

Animal species : Wistar albino rats

Age : 6-8 weeks

Weight : 150-200 gms

Gender : Both male and female

Number of animals : 24

Acclimatization period : 7 Days

P a g e | 73

ANIMAL SOURCE:

Test animals were obtained from The King Institute, Chennai, and kept at

animal house, National Institute of Siddha, Chennai. All the animals were kept under

standard environmental condition (22 ± 3˚c) .The animals had free access to water and

standard pellet diet (Sai Meera foods pvt.ltd, Bangalore). The principles of laboratory

animal care were followed.

RANDOMIZATION, NUMBERING AND GROUPING OF ANIMAL

The animals were randomly divided into four groups. Each group consist of 6

animals (3 per sex in each group) first group was kept as control and remaining

groups as test were treated with test drug in different doses. The animals were allowed

for an acclimatization period of 7 days to laboratory conditions prior to the initiation

of treatment. Each animal was marked with picric acid. The females were nulliparous

and non pregnant.

IDENTIFICATION OF ANIMAL:

By cage number and individual marking on the fur of each animals with picric

acid.

HOUSING & ENVIRONMENT:

The animals were housed in polypropylene cages provided with bedding of

husk. Dark and light cycle each of 12 hours was maintained.

S.No Groups No of Rats

1 Vehicle control 6 (3 male, 3 female)

2 1X Therapeutic dose (0.1296 mg/animal) 6 (3 male, 3 female)

3 5X Therapeutic dose (0.648 mg/animal) 6 (3 male, 3 female)

4 10X Therapeutic dose (1.296 mg/animal) 6 (3 male, 3 female)

P a g e | 74

ADMINISTRATION PERIOD:

The long term toxicity study was carried out for one month because the

clinical duration of human consumption of test drug is 8 days.

DOSE SELECTION:

The long term toxicity study was carried out at different dose level 0.1296

mg/animal, 0.648 mg/animal, 1.296 mg/animal. The selected doses were calculated

according to body weight, and surface area of rat. The human therapeutic dose of

Poora Parpam is 12mg/day.

PREPARATION AND ADMINISTRATION OF DOSE

Poora Parpam was suspended in aqueous tween 80 solution (10%). It was

administered to groups at dose levels of X therapeutic dose (0.1296 mg/animal), 5X

therapeutic dose (0.648 mg/animal) and 10X therapeutic dose (1.296 mg/animal). The

control animals were administered vehicle only. Administration was given orally

using an oral gavage once in daily for 30 days.

OBSERVATION:

Experimental animals were kept under observation throughout the course of

the study and recorded for the following:

MORTALITY:

All animal were observed twice daily for mortality during the entire study

period.

BODY WEIGHT:

Weight of each rat was recorded on first day and at weekly intervals

throughout the course of study and at the end of the study. From the data mean body

weights and percent body gain were calculated.

P a g e | 75

FOOD AND WATER INTAKE:

The quantity of food and water consumed by animals of different doses was

recorded at weekly intervals. Food and water consumed per animal was calculated for

control and the treated dose groups.

LABORATORY INVESTIGATIONS:

Collection of blood:

At the end of study the blood samples were collected by cardiac puncture

using as syringe. The collected blood samples were kept in vaccutainer blood samples

were centrifuged at 3000 rpm for 10 minutes and collected serum was used for

laboratory investigations.

Hematological parameter:

Hematological parameters like Hb, Total RBC, Total WBC, DC, ESR were

analyzed.

Biochemical parameter:

Blood Glucose, Lipid profile, T.bilirubin, D. bilirubin, I. bilirubin, SGOT,

SGPT, ALP, T.Protein, albumin, globulin, Urea, Creatinine, Uric acid, Calcium and

Potassium were determined.

NECROPSY:

All the animals were sacrificed at end of the study under ether anesthesia.

Necropsy of all animals was carried out and the morphological changes of organs

including liver, kidneys, brain, heart, and lungs were noted.

P a g e | 76

HISTOPATHOLOGY:

Tissue samples of organs from control and treated animals were preserved in

10% formalin for preparation of sections using microtome. The organs liver, kidneys,

spleen, brain, heart and lungs of the animals were preserved. They were subjected to

histopathological examination.

The organ pieces (3 to 5 micron) were fixed in 10% formalin for 24 hours and

washed in running water for 24 hours. Samples were dehydrated in tissue processor

and then cleaned in benzene to remove absolute alcohol. Embedding was done by

passing the cleared sample through three cups containing molten paraffin at 50˚c and

then a cubical block of paraffin made with L moulds, followed by microtome slicing

and mounting on the slides. The slides were stained with Haematoxylin – Eosin stain.

STATISTICAL ANALYSIS:

Findings such as clinical sings of intoxication, body weight changes, food and

water consumption, hematology and biochemical parameters were subjected to one-

way ANOVA followed by Dunnett’s “t” test using a computer software programme-

INSTAT V3 version.

P a g e | 77

CHAPTER V

5. RESULTS

PHYSICO CHEMICAL ANALYSIS

Colour characters of Poora Parpam.

Table.3.

Physico chemical properties of Poora Parpam.

S No

Nature of the drug

Under ordinary light

Under ultra violet light

1.

Powdered material

White

White

S No.

Parameters

Values obtained (%w/w)

1.

Total ash value

9.66

2.

Acid insoluble ash

0.92

3.

Water soluble ash

5.1

4.

Moisture content

8.58

5.

Foreign organic matter

8.3

6.

Alcohol soluble extractive

6.2

7.

Water soluble extractive

10.20

8.

Loss on drying at 105˚C

7.2

P a g e | 78

Colour, nature and percent yields of extracts of Poora Parpam.

QUALITATIVE ANALYSIS

CHEMICAL ANALYSIS

The qualitative analysis of unpurified Pooram show the presence of

Mercury

Phosphate

Iron

Magnesium

The qualitative analysis of purified Pooram show the presence of

Mercury

Phosphate

Iron

Magnesium

The qualitative analysis of Pooram Parpam show the presence of

Mercury Starch

Phosphate Tannic acid

Iron

Magnesium

S.No.

Extract

Solvents

Colour

Nature

%Yield(w/w)

pH

1.

Water

White

Solid

45

7.2-7.5

P a g e | 79

QUANTITATIVE ANALYSIS

HR SEM Analysis - Determination of particle size of Pooram Parpam

Fig.4. HR SEM - POORA PARPAM

1. Particle size is 1 to 10 µ (micron).

2. Cumulative distribution of the particles with bulk density is seen.

3. Surface is smooth in nature.

P a g e | 80

INDUCTIVELY COUPLED PLASMA OPTICAL EMISSION SPECTROMETRY

Table. 4.

S.no.

Elements

Wavelength

in nm

Unpurified

Pooram

(ppm)

Purified

Pooram

(ppm)

Poora

Parpam

(ppm)

1.

Arsenic

As193.696

BDL*

BDL*

BDL*

2.

Calcium

Ca 317.933

18.654

16.425

15.856

3.

Cadmium

Cd 226.502

BDL*

BDL*

BDL*

4.

Mercury

Hg 253.652

204.175

98.251

3.187

5.

Iron

Fe 238.204

1.856

1.658

1.356

6.

Potassium

K 766.490

47.853

46.522

45.426

7.

Sodium

Na 589.592

20.689

19.853

17.952

8.

Phosphate

P 213.617

11.751

10.856

9.784

9.

Lead

Pb 230.204

BDL*

BDL*

BDL*

* BDL = below detection limit, ppm – Parts per million

P a g e | 81

TOXICITY STUDY

Acute oral toxicity study

All the data´s were summarized in the form of table (5) showing the

animals behavioral signs in control and test groups. There was no mortality at

the dose level of 0.216 mg/ animal. (10 times more than the therapeutic dose).

Long term toxicity study

Clinical signs

No abnormal behavioral signs were observed during the study period.

Mortality

The test drug Poora Parpam did not cause any mortality in X (0.1296

mg/animal), 5X (0.648 mg/animal) and 10X (1.296 mg/animal) dose levels

and were considered as safe dose levels.

Body weight

Both control and test dose groups exhibited normal body weight

throughout the study period. Table. (6).

Food consumption

No difference in food intake of control and test group animals

observed during the period of study. Table. (8)

Water intake

No difference in water intake of control and test group animals

observed during the period of study. Table. (7).

P a g e | 82

Hematological investigations

The results of hematological investigation conducted at the end of the

study, test groups revealed no significant changes in values of different

parameters, when compared with control group. The Lymphocyte count was

slightly elevated in test groups, but statistically not significant when compared

with control group. Table. (9)

Biochemical investigations

Biochemical investigations were conducted at the end of the study and

the results were recorded. In test groups there was no significant elevation in

the levels of biochemical parameters, when compared with the control group.

And the values obtained were within normal biological limits. Table. (10 - 12)

Histopathology

Gross pathological examination of animals doesn’t reveal any

abnormalities in control and test groups. The histopathological study of the

organs such as heart, lungs, kidney, brain and liver was normal in control, X,

and 5X groups. In 10 X group the brain shows mild edema in cerebral cortex

with separated glial cells. The lung shows bronchioles, alveoli, interstium

shows dilated, congested blood vessels. Focal lymphoid aggregations were

present. The kidney shows congested glomeruli. Tubules appear normal.

Interstium shows congested blood vessels with area of hemorrhage. The heart

shows normal myocardial fibers with congested blood vessels and

hemorrhage.

P a g e | 83

HISTOPATHOLOGICAL REPORT

BRAIN

Plate .a. CONTROL

Sections show normal structure of cerebral parenchyma, cerebellum.

Plate .b. X group

Section shows normal structure of cerebral parenchyma, cerebellum.

Plate .c. 5X group

Section shows normal structure of cerebral parenchyma, cerebellum.

Plate .d. 10X group

Section of the brain shows mild edema in cerebral cortex with separated glial cells .

P a g e | 84

BRAIN

Plate.a.Control group Plate.b. X group

Plate.c. 5X group Plate.d. 10X group

Plate.e. 10X group - h igh power – Showing edematous cerebral cortex with separated glial cells.

P a g e | 85

HEART

Plate .a. CONTROL

Sections show normal myocardial fibers and blood vessels.

Plate .b. X group

Section shows normal myocardial fibers and blood vessels.

Plate .c. 5X group

Section shows normal myocardial fibers and blood vessels

Plate .d. 1OX group

Section of the heart shows myocardial fibers with congested blood vessels and

hemorrhage.

P a g e | 86

HEART

Plate.a.CONTROL Plate.b.X group

Plate.c. 5X group Plate.d.10X group showing normal myocard ial

fibers with Congested blood vessels and

haemorrhage.

P a g e | 87

KIDNEY

Plate .a. CONTROL

Specimen shows renal parenchyma with normal appearing cortex and medulla.

Plate .b. X group

Specimen shows renal parenchyma with normal appearing cortex and medulla.

Plate .c. 5X group

Specimen shows renal parenchyma with normal appearing cortex and medulla.

Plate .d. 10X group

Section from the kidney shows congested glomeruli. Tubules appear normal.

Interstitium shows congested vessels with area of hemorrhage.

P a g e | 88

KIDNEY

Plate.a. CONTROL Plate.b. X group

Plate.c. 5X group Plate.d. 10X group showing congested

blood vessels an haemorrhage.

Plate.e.10X group - high power - showing congested glomeruli.

P a g e | 89

LIVER

Plate .a. CONTROL

Section from the liver shows normal appearing central veins with radiating

hepatocytes, sinusoids, kuffer cells and portal triad.

Plate .b. X group

Section from the liver shows normal appearing central veins with radiating

hepatocytes, sinusoids, kuffer cells and portal triad.

Plate .c. 5X group

Section from the liver shows normal appearing central veins with radiating

hepatocytes, sinusoids, kuffer cells and portal triad.

Plate .d. 10X group

Section from the liver shows normal appearing central veins with radiating

hepatocytes, sinusoids, kuffer cells and portal triad.

P a g e | 90

LIVER

Plate.a. CONTROL Plate.b. X group

Plate.c.5X group Plate.d.10X group

Plate.e.10X group - high power Plate.f.10X group showing normal

portal triad with radiating sinusoids

P a g e | 91

LUNG

Plate .a. CONTROL

Section from the lung shows normal appearing bronchioles, alveoli, interstitium and

blood vessels

Plate .b. X group

Section from the lung shows normal appearing bronchioles, alveoli, interstitium and

blood vessels

Plate .c. 5X group

Section from the lung shows normal appearing bronchioles, alveoli, interstitium and

blood vessels

Plate .d. 10X group

Section of the lung shows bronchioles, alveoli, interstitium with dilated, congested

blood vessels. Focal lymphoid aggregations are present.

P a g e | 92

LUNG

Plate.a. CONTROL Plate.b. X group

Plate.c .5X group Plate.d. 10X group showing bronchioles ,

alveoli, interstium shows dilated,

congested blood vessels. Focal

lymphoid aggregations were present.

P a g e | 93

Table.5. Behavioral signs of Acute toxicity study in S wiss albino mice treated with Poora Parpam

1. A lertness 2. Aggressiveness 3. Passivity 4. Grooming 5. Gripping strength 6. Touch responses

7.Restlessness 8. Tremors 9. Convulsion 10. Pain response 11.Defaecation 12. Pinna reflex13.Corneal

reflex 14.Pupillary size15. Lacrimat ion 16. Salivation 17. Urination 18. Writhing 19.Skin colour 20.

Mortality

2. + Presence of activity, N - Normal,

- Absence of activity A - Absent

Table.6. Body wt (g) of Wister rats in long term toxicity study treated with Poora Parpam

NS- Not Significant, *(p >0.05), n = 6 values are mean ± S.D (One way Anova followed by

Dunnett’s test)

No.

Treatment

group

1

2

3

4

5

6

7

8

9

10

11

12

13

14

15

16

17

18

19

20

1.

Control

+

–

–

+

+

+

–

–

–

+

–

+

+

+

–

–

–

–

N

A

2.

Acute

Dose

(216mg/

animal)

+

–

–

+

+

+

+

–

–

+

+

+

+

+

–

–

–

–

N

A

DOS E

(mg/animal)

DAYS

1 7 14 21 30

CONTROL

117.55 ± 3.57

113.60 ± 5.83

122.05 ± 3.54

122.50 ± 4.54

130.15 ± 3.24

X - GROUP

112.33 ± 2.36

119.01 ± 2.66

120.48 ± 3.44

121.60 ± 4.15

132.13 ± 4.11

5X -

GROUP

115.47 ± 4.12

118.23 ± 3.44

119.86 ± 3.89

130.77 ± 3.33

130.19 ± 4.53

10 X -

GROUP

119.49 ± 3.89

120.22 ± 4.25

121.76 ± 3.09

130.47 ± 5.55

135.13 ± 4.58

P value (p)*

NS

NS

NS

NS

NS

P a g e | 94

Table.7. Water (ml/day) of Wister rats in long term toxicity study treated with Poora Parpam

DOS E

(mg/animal)

DAYS

1

7

14

21

30

CONTROL

25.55 ± 2.54

23.60 ± 2.83

33.05 ± 3.59

34.5 ± 4.57

30.15 ± 3.24

X - GROUP

22.33 ± 3.44

31.01 ± 2.66

34.48 ± 4.17

42.6 ± 4.15

37.23 ± 3.44

5X -

GROUP

35.47 ± 3.89

38.23 ± 3.36

31.86 ± 3.89

36.77 ± 3.33

38.19 ± 4.53

10 X -

GROUP

33.49 ± 4.58

40.22 ± 4.25

36.76 ± 3.09

44.47 ± 5.55

41.13 ± 4.12

P value (p)*

NS

NS

NS

NS

NS

N.S- Not Significant, *(p >0.05), n = 6 values are mean ± S.D (One way Anova followed by

Dunnett’s test)

Table.8. Food (g/kg) intake of Wister rats in long term toxicity study treated with Poora Parpam

DOS E

(mg/animal)

DAYS

1

7

14

21

30

CONTROL

27.55 ± 3.24

33.60 ± 3.55

39.05 ± 3.25

41.5 ± 2.05

43.15 ± 4.76

X - GROUP

26.33 ±4.15

39.01 ± 2.66

34.48 ± 3.44

38.6 ± 3.89

41.32 ± 4.11

5X GROUP

31.47 ± 4.12

35.23 ± 3.44

32.86 ± 3.89

35.77 ± 2.36

44.19 ± 4.53

10 X

GROUP

28.49 ± 3.33

31.22 ± 4.25

35.76 ± 3.09

43.47 ± 5.58

45.13 ± 4.58

P value (p)*

NS

NS

NS

NS

NS

N.S- Not Significant,* (p >0.05),n = 6 values are mean ± S.D (One way Anova followed by

Dunnett’s test)

P a g e | 95

Table.9. Haematological parameters of Wister rats in long term toxicity study treated

with Poora Parpam

NS- Not Significant,* (p >0.05) ** S – Significant (p<0.05) n = 6 values are mean ± S.D

(One way Anova followed by Dunnett’s test)

Table.10. Biochemical Parameters of Wister rats in long term toxicity study treated with

Poora Parpam

BIOCHEMICAL

PARAMETERS

CONTROL

X GROUP

5X

GROUP

10X

GROUP

P Value

(p)*

GLUCOS E (R) (mg/dl)

100 ± 40

112 ±18

116 ± 37

90 ± 25

N.S

T.CHOLOSTEROL(mg/dl)

68 ± 7

70 ±10

67 ± 5

71 ± 4

N.S

HDL(mg/dl)

20 ± 4

23 ± 5

22 ± 5

24 ± 2

N.S

LDL(mg/dl)

22 ± 7

22 ± 8

20 ± 6

22 ± 6

N.S

VLDL(mg/dl)

27 ± 4

25 ± 4

25 ± 6

25 ± 4

N.S

TRIGLY(mg/dl)

133 ± 20

127 ± 17

124 ± 28

129 ±18

N.S

NS- Not Significant,* (p >0.05), n = 6 values are mean ± S.D (One way Anova followed by

Dunnett’s test)

Category

Haemoglobin

(g/dl)

Total -WBC

(cells/cu.mm)

Diff.Count

Neutrophil

( % )

Diff.Count

Lymphocyte

( % )

Total - RBC

(cells/cu.mm)

CONTROL

13.4 ± 0.6

4850 ± 495

40 ± 31

55 ± 24

4.2 ± 0.1

X

15.5 ± 2.2

5833 ± 2017

21 ± 9

78 ± 9

5.8 ± 1.2

5X

16.4 ± 3.5

4717 ± 1598

24 ± 8

76 ± 8

5.1 ± 1.0

10X

14.0 ± 2.5

4750 ± 1435

24 ± 8

77 ± 8

5.5 ± 1.3

P value (p)* NS NS NS NS, **

5X vs. X -S NS

P a g e | 96

Table.11. Renal function test of Wister rats in long term toxicity study treated with

Poora Parpam

PARAMETERS CONTROL X - GROUP 5X -

GROUP

10X -

GROUP

P Value

(p)*

UREA (mg/dl) 28 ± 16 26 ± 3 28±7 30 ± 6

N.S

CREATININE

(mg/dl) 0.76 ± 0.25 0.69 ± 0.10 0.66 ± 0.09 0.72 ± 0.12

N.S

URIC ACID(mg/dl) 2.51 ± 0.20 2.57 ± 0.36 2.38 ± 0.50 2.43 ± 0.37

N.S

CALCIUM(mg/dl) 8.8 ± 0.8 8.9 ± 0.8 8.7 ± 0.6 9.1 ± 0.6

N.S

POTASSIUM

(mg/dl) 2.7 ± 0.4 2.7 ± 1 2.6 ± 0.5 3.1 ± 0.6

N.S

NS- Not Significant,* (p >0.05) , n = 6 values are mean ± S.D (One way Anova followed by Dunnett’s

test)

Table.12. Liver Function Test of Wister rats in long term toxicity study treated with

Poora Parpam

PARAMETERS CONTROL X - GROUP 5X -

GROUP

10X -

GROUP

P Value

(p)*

T.BILIRUBIN(mg/dl)

0.9 ± 0.1

0.7 ± 0.1

0.7 ± 0.2

0.7 ± 0.2

N.S

D.BILIRUBIN(mg/dl)

0.3 ± 0.0

0.3 ± 0.1

0.3 ± 0.1

0.3 ± 0.1

N.S

I. BILIRUBIN(mg/dl)

0.6 ± 0.1

0.4 ± 0.1

0.4 ± 0.2

0.4 ± 0.1

N.S

SGOT(U/dl)

67 ± 25

70 ± 23

58 ± 16

57 ± 16

N.S

SGPT(U/dl)

77 ± 31

86 ± 27

66 ± 23

62 ± 22

N.S

ALP(U/dl)

132 ± 6

142 ± 25

138 ± 8

134 ± 20

N.S

T.PROTEIN(mg/dl)

7.0 ± 0.9

6.7 ± 0.3

6.8 ± 0.5

7.1 ± 0.4

N.S

ALBUMIN(mg/dl)

3.3 ± 0.6

3.4 ± 0.4

3.2 ± 0.3

3.3 ± 0.2

N.S

GLOBULIN(mg/dl)

3.7 ± 0.3

3.3 ± 0.2

3.6 ± 0.5

3.7 ± 0.3

N.S

NS- Not Significant,* (p >0.05), n = 6 values are mean ± S.D (One way Anova followed by Dunnett’s

test)

P a g e | 97

Chart.1. Mean arithmetic values of Control and test group animals

Note: The lipid levels compared to control group were equal in test groups

Chart.2. Mean arithmetic values of Control and test group animals

Note: The LFT levels compared to control group were equal in test groups

P a g e | 98

Chart.3. Mean arithmetic values of Control and test group animals

Note: The LFT levels compared to control group were equal in test groups

Chart.4. Mean arithmetic values of Control and test group animals

Note: The RFT levels compared to control group were equal in test group

P a g e | 99

Chart.5.Mean arithmetic values of Control and test group animals

Note: The lymphocyte levels compared to control group, increased in test groups but not significant statistically

Chart.6. Mean arithmetic values of Control and test group animals

Note: The T.WBC levels compared to control group were increased in test groups (X)

P a g e | 100

Chart.7.Mean arithmetic values of Control and test group animals

Note: The T.RBC levels compared to control group were increased in test groups

Chart.8. Mean arithmetic values of Control and test group animals

Note: The Glucose and Urea levels compared to control group, varied in test groups

but not significant statistically

P a g e | 101

CHAPTER VI

6. DISCUSSION

In this study Poora Parpam was subjected to qualitative, quantitative and

toxicity studies. Qualitative analysis includes chemical analysis of unpurified, purified

and Pooram Parpam. Quantitative analysis includes ICP OES, HR SEM. And then

finally both acute and long term toxicity study were carried out in rodents as per

WHO guidelines.

Qualitative Analysis of unpurified and purified Pooram sample indicated the

presence of mercury, phosphate, iron and magnesium. Analysis of the Poora Parpam

showed the presence of mercury, phosphate, iron, magnesium, starch and tannic acid.

This reveals that there is no change in the unpurified and purified sample. But

in prepared medicine there is addition of starch and tannic acid compared to the

purified and unpurified Pooram. Because the Pooram Parpam was prepared with the

madar latex juice and onion juice, to increase the efficacy. Hence the presence of

starch and tannic acid. (Table 1, 2)

In the Physico Chemical Analysis the pH of Pooram Parpam was 7.2 -7.5. It

denotes weak alkalinity. Thus, on oral intake it will not cause any strong alkali or acid

like irritation to the gastrointestinal tract. The loss on drying at 105ºc was only 7.2%

w/w; hence the drug will not loose much of its volume on exposure to atmospheric air

at room temperature. Solubility of Poora Parpam was found to be increased compared

to the purified and unpurified Pooram. Hence there is possibility of increased

absorption. (Table.3.)

HR SEM analysis of Poora Parpam reveals the particle size as 1-10 µ

(micron). The particles were homogenously distributed in the Parpam. And smooth

surface of the particles enable its easy for absorption in the gastro intestinal tract.

Hence the drug will have increased bioavailability. (Fig.4.)

P a g e | 102

In ICP-OES study, heavy metals like As, Pb, Cd were found below detection

limit in unpurified and purified Pooram. Mercury, calcium, iron, potassium,

phosphorous, sodium levels were decreased in its level from unpurified to purified

Pooram.

There was a drastic reduction in the level of mercury in unpurified Pooram

(204.175) to purified Pooram (98.251). Mercury was detected in level in the finally

prepared medicine Poora Parpam as (3.187). This indicates that all heavy metals in

Poora Parpam were within normal range and near to permissible limit. So that Poora

Parpam is safe for human consumption. (Table.4.)

In Acute toxicity study period there were no abnormal signs developed in

Swiss albino mice at 10 times more than the therapeutic dose level (0.216 mg/animal)

within 24 hrs. No mortality and reduction in body weight of animals were observed in

the study period. (Table.5.)

At the end of Long term toxicity study animals were sacrificed and blood

samples were collected and investigated. The organs were collected and sent for

histopathology study. All the reports were statistically calculated. There were no

significant changes in hematological parameters, biochemical investigations, body

weight, food and water intake. There was a marked increase in lymphocyte count in

test groups, but compared to control group, it is not statistically significant.

(Table.6-12).

The histopathological study on the organs such as heart, lungs, kidney, brain

and liver was normal in control, X, and 5 X groups. In 10 X group, the brain shows

mild edema in cerebral cortex with separated glial cells. The lung shows focal

lymphoid aggregations, congested blood vessels and interstitium was dilated. The

kidney shows congested glomeruli and interstitium shows congested blood vessels

with area of hemorrhage. The heart shows congested blood vessels and hemorrhage.

P a g e | 103

CHAPTER VII

7. SUMMERY

Siddha system is the ancient traditional system followed in South India mainly

in Tamilnadu. It incorporates many herbal, mineral and metal based medicines.

Pooram is mentioned under minerals and Poora Parpam is one of the notable medicine

in the treatment of arthritis, syphilis and jaundice. Poora Parpam taken from the

literature Veera Maamunivar Vaagada Thiratu in this study also indicated for the

treatment of arthritis.

The raw drug Pooram was procured from country drug shop and authenticated

at Siddha Central Research Institute. Pooram was purified and the medicine was

prepared as mentioned in the Siddha literature.

Chemical analysis of unpurified and purified Pooram sample indicates the

presence of mercury, phosphate, iron and magnesium. Analysis of the Poora Parpam

confirmed the presence of mercury, phosphate, iron, magnesium, starch and tannic

acid.

In physico - chemical analysis of Pooram Parpam the pH was found to be 7.2

-7.5 and the loss on drying at 105ºc was 7.2% w/w.

After HR SEM analysis, the particle size of the Poora Parpam was analyzed as

1-10 µ (micron).

In ICP-OES study, heavy metals like As, Pb, Cd were found below detection

limit in unpurified and purified Pooram. Mercury, calcium, iron, potassium,

phosphorous, sodium levels were decreased in its level from unpurified to purified

Pooram. Mercury was detected in minimal level in the finally prepared medicine

Poora Parpam as (3.187).

P a g e | 104

The toxicological evaluations were conducted as per WHO guidelines for

safety evaluation of Poora Parpam.

In Acute toxicity study there were no abnormal signs developed in Swiss

albino mice at 10 times more than the therapeutic dose level (0.216 mg/animal) within

24 hrs. At the end of the study no mortality and reduction in body weight o f control

and test group animals were observed.

In Long term toxicity study there were no significant changes in behavioral

signs, hematological parameters, biochemical investigation, body weight, food and

water intake. The lymphocyte count was increased in test groups, but compared with

control group it is not statistically significant. The histopathological study on the

organs such as heart, lungs, kidney, brain and liver was normal in X and 5 X groups

compared with control group. In 10 X group, only the brain showed mild edema with

separated glial cells and the other organs showed no abnormal histological variation.

P a g e | 105

CHAPTER VIII

8. CONCLUSION

As the result of this study, it has been concluded that

The decrease in heavy metal content of the prepared medicine when compared

to raw drug analyzed by sophisticated instrument reveals the potential background of

Siddhars knowledge in the field of metals, minerals and herbal usage in medicine

preparation. Mainly the mercurial content of the prepared medicine showed drastic

reduction when compared to that of unpurified raw drug (204.175 to 3.187).

The acute and long term toxicity of Poora Parpam is found to be less toxic and

the therapeutic dose level mentioned in the literature is a safer dose for human

consumption.

Hence further studies on Poora Parpam are to be conducted for its scientific

validation and global acceptance.

P a g e | 106

BIBLIOGRAPHY

1. Dr.R.Thiagarajan L.I.M, Gunapadam Thathu Jeeva Vaguppu 4th edition 2004,

Published by Indian Medicine & Homeopathy Dept. Chennai – 106.

2. S.P.Ramachandran, Veerama munivar vagadathiratu,Thamarai Noolagam,

Chennai 26.

3. Dr.K.S.Murugesa mudhaliyar,H.P.I.M, Gunapadam Mooligai,7th edition 2003,

Published by Indian Medicine & Homeopathy Dept. Chennai – 106.

4. Hakkim P.M.Abdulla Sahib, Anuboga Vaidhya Navaneetham, part-4,

published by Thamarai Noolagam,Chennai,26.

5. S.P.Ramachandran, Yaahoobu soonagadam, Thamarai noolagam, Chennai -26

6. Dr.K.S.Murugesa Mudhaliyar, Nanju murivu nool, 3rd edition, 1998 revised by

Dr.Pongurusiromani, Dept. of Indian Medicine & Homeopathy,Chennai – 106.

7. S.P.Ramachandran, Agathiyar Silaraikovai, 1996, Thamarai noolagam,

Chennai -26.

8. Dr.Kandaswamy mudhaliyar, Aathma Ratchamirthamennum Vaidhya

Sarasangiragam, 1st edition 1993,Rathna Nayakar & sons, Thirumagal vilasa

printers, Chennai – 79.

9. Sarabenthirar Vaithya Muraigal published by Saraswathi Mahal Library,

Tanjore – 09.

10. Dr.S.Arangarajan, B.I.M, Agathiyar Attavanai Vagadam 1st edition 1991 by

Saraswathi Mahal, Tanjore – 07.

11. Dr.Kannusamy pillai, Pathartha guna sinthamini, Ratna Nayakar & sons,

Chennai – 79.

12. S.P.Ramachandran, Theriyar Vaidhya Kaviyam 50, 2nd 2000, Thamarari

Noolagam, Chennai -26.

13. S.P.Ramachandran, Agathiyar Vaidhya Kaviyam 1500, Thamara i noolagam,

Chennai 26.

14. Dr.Kannusamy Pillai, Kannusamy Parambarai Vaidhyam 2006- Rathna

nayakar & sons, Chennai – 79.

15. S.P.Ramachandran, Gorakkar Chandra Regai, Pulipaani Jaala thiratu,

Thamarai Noolagam , Chennai 26.

P a g e | 107

16. Sirumanavoor Munnuswamy Mudhaliyar, Visha Vaidhya chindamani, Rathna

Nayakar & sons, Thirumagal vilasa printers, Chennai – 79.

17. S.P.Ramachandran,Uyirkakkum Siddha Maruthuvam, Thamarai Noolagam,

Chennai- 26.

18. Dr. C.A. Franklin MD – Modi’s Medical Jurisprudence & Toxicology 21st

edition

19. The Wealth of India, Council of Scientific and Industrial Research New Delhi.

20. Dr.K.M.Nadkarani, Indian Materia Medica, 3rd edition, Bombay popular

prakashan.

21. Dr.T.V.Sambasivam pillai, Dictionary of Medicine, Chemistry, Botany &

Allied Science (based on Indian Medical Sciences) Volume I- V.

22. Dr.K.S.Narayana Reddy, The Essentials of Forensic Medicine and Toxiclogy,

29th edition Published by K.Sugana Devi, Hyderabad – 36.

23. Thomas W. Clarkson, The Toxicology of Mercury and Its Chemical

Compounds, Critical Reviews in Toxicology, 36:609–662, 2006 ISSN: 1040 -

8444 print / 1547-6898

24. Adak M, Gupta J.K, Evaluation of anti-inflammatory activity of Calotropis

gigantea (AKANDA) in various biological system, Nepal Med Coll J. 2006

Sep, 8(3):156-61

25. Himanshu Joshi, Gururaja M.P and Divya Suares, Calotropis gigantea R. Br.

(Asclepiadaceae), International Journal of Pharmaceutical Research 2011,

Volume 3, Issue 1, 10-14, ISSN 0975-2366.

26. Kumar VL, Basu N, Anti-inflammatory activity of the latex of Calotropis

procera, J.Ethnopharmacol, 1994 Oct, 44(2):123-5.

Evan A. Ross, Medicinal Plants of the World, Volume 2, and published by

27. Humana Press, Totowa, New Jersey. (2001),

K.Vamshi Sharath Nath, Rao KNV, David Banji, Sandhya S, Sudhakar K,

Saikumar P, Sudha P, Chaitanya R.K, A comprehensive review on Allium

cepa, Journal of Advanced Pharmaceutical Research 2010, 1(2), 94-100,

ISSN: 2229‐3787.

P a g e | 108

28. Chitra Shenoy , M B Patil , Ravi Kumar And Swati Patil , Preliminary

Phytochemical Investigation and Wound Healing Activity of Allium Cepa

Linn (Liliaceous), International Journal of Pharmacy and Pharmaceutical

Sciences, Vol. 2, Issue 2, July Sep.2009

29. Palache, C., H. Berman, and C. Frondel (1951) Dana’s system of mineralogy,

(7th edition), V, II, 25–28.

30. Calos, N.J, C.H.L. Kennard, and R.L. Davis (1989), The structure of

calomel, Hg2Cl2 derived from neutron powder data. Zeist. Krista, 187,

305– 307. (3)(1976) NBS Mono, 25, 13, 30.

31. Http://cartwright.chem.ox.ac.uk/hsci/chemicals/mercury_I_chloride.html

32. Kegley, S.E., Hill, B.R., Orme S., Choi A.H., PAN Pesticide Database,

Pesticide Action Network, North America (San Francisco, CA, 2010),

http:www.pesticideinfo.org.

33. Narendranalwaya,Gauravpokharna,Lokeshdeb,Naveen Kumarjain, Wound

Healing Activity of Latex of Calotropis Gigantea, International Journal of

Pharmacy and Pharmaceutical Sciences, Vol. 1, Issue 1, July -Sep. 2009