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c   c c
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ccccc c

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cc c !" cc


"#"c !cc$%"cc&c
c'$("%c "%)*c **"+",c ' "c
ccccccccccccc%-$#.+#*) #ccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccc c
cc  !" c%cc
"#"c !c ) / * +'c
c "%)*c **"+",c ' "c
c

cc *c "cc)& *c


c"#"c !cc$%"cc&c
ccccccccccccc'$("%c "%)*c **"+",c ' "c
cccccccccccc&0"&+&12.+#*) #c

 
c
c

3c

 :  c   [copper ash] is used in treating various disorders like   c


[fever], 

  [skindisorders] and based on its antipyretic, anti parasitic and
antileprotic properties mentioned in our classics, the present study was undertaken to
evaluate antibacterial role of   c  on gram positive and gram negative bacteria.c

  
cc :c  and other raw drugs such as   [mercury]and
 
[sulphur] was purified and  c  was subjected to    c[incineration]
process according to the procedures mentioned in classics.
MIC and MBD of the prepared sample was determined by broth dilution method by
following NCCLS guidelines.

 
4  c  c had an antibacterial effect against both gram positive and gram
negative bacteria. Minimum inhibition concentration [MIC] and minimum bactericidal dose
[MBD] was estimated to be 2.5mg/ml of nutrient broth for E coli and between 1.25mg/ml for
staphylococcus


 : the exact therapeutic dose detection needs further detailed analysis.

 c:

c MIC ±Minimum inhibitory concentration


c MBD ± Minimum bactericidal dose
c  c  ±[ Copperash ]
c ×   [Antipyretic]
c ’   c[ Anti parasitic and Antibacterial]
c ’  c[ cures skin disorders or anti leprotic]

c
ÿ 
c
c

 4c

 has been used in preparing various medicines for thousands of years.Our ancient
classics mentions the therapeutic use of copper in treating various disorders such as
   c [conjunctivitis],
   c [antihelminthic & antiparasitic],  
[cholera],
variousc  c[fever], 

 c[skin disorders] etc.

The following Historical review on copper shows it has antibacterial effect

Xc The first medical use of copper found in papyrus the Egyptian medical text records
use of copper in sterilizing chest wounds.
Xc P  c mentioned preparation of  
[needle] out ofc  in operation
of cataract.
Xc In Hippocratic collection copper was recommended in the treatment of leg ulcers
associated with varicose veins.
Xc Àreeks used dry powder composed of copper oxide and copper sulphate on the
wound.
Xc Inorganic copper preparations were found to be effective in treating eczema,
impetigo ,tubercular infections etc.

Thus based on its use on certain bacterial infections since ancient times and important
properties such as    c c
   c and
  mentioned in our classics
present study on   c   was undertaken to prove its efficacy as an antibacterial
agent.

 cc   4c

Xc To carry out Purification of the raw drugs ±   [mercury],  


c[sulphur]
and  [copper].
Xc To prepare  c  [COPPER ASH]
Xc To find the Antibacterial action of the prepared sample and
Xc To determine the MIC & MBD of prepared sample against gram positive bacteria and
gram negative bacteria.

  
cc 4c

Raw materials were collected from genuine sources and subjected initially for the purification
process.

PARADA SHODHANAc5  c  6c718c

REFERENCE: "  c  c   cc


METHOD:  c   method
EQUIPMENTS:cc’    c c   c  
INÀREDIENTS: P c   ± 500gc
  c   [aloe vera juice]± Q.S,
   [turmeric] ± 500g
* 
c
c

  DcMercury and Turmeric taken in known quantity was triturated well till the
complete mercury is assimilated into Turmeric then aloe vera juice was added and 


[small thin plates] were prepared with it . 

 dried and placed into  c  
  [equipment to procure mercury] and subjected for heating for 6 hours. After self
cooling mercury was collected and washed thoroughly in hot water for 3-5 times.
/"( 4cc

P"P Pc !"#"!c$# PPc P" !"c$# PPPc


îEIÀHT 500g 478g
APPEARANCE Dull shine, Lustrous, clear and shiny

A] PURIFIED MERCURY B] PURIFIED SULPHUR

ÀANDHAKA SHODHANA5  c c


c67198c
c
REFERENCE: "  c  c   c%&%c
METHOD:   c
EQUIPMENTS: Mud pot, $  c 
INÀREDIENTS: c 
± 500g,  ± 2litres
PROCEDURE: mud pot was filled with  andc 
was placed on cloth tied
over its mouth and covered with   c  c'c   [cow dung cakes] placed above in
sufficient quantity and ignited after self coolingc 
collected in the milk was collected
and washed in hot water.

OBSERVATIONS:

P P’Pc !"#"!c$# PPcc P" !"c$# PPc


îEIÀHT 500g 496g
APPEARANCE Yellow, Crystalline Small Àlobular and lustrous
c
c
c
c
c
c
c
è 
c
c

AMRAc  
cc  c  4c7:8c
REFERENCE: "  c  c   5/29
EQUIPMENT: Steel vessel

INÀREDIENTS:
   c   [general purification] -Tamra -1kg,    c 
 c  c    ,

 
 , in sufficient quantity
(   cc  c[specific purification]c)c   c  c  c  c   cand
   c   c c  c[lemon juice] cin quantity sufficient
PROCEDURE:    c   was done by heating and dipping for 7 times in each
above said liquid.and (   c   was done by applying paste of saindhava and
 c  on   c  cheating and dipping it inc  c  this process was done for
8 times

C] COPPER D] PURIFIED COPPER

OBSERVATIONS : copper general purification and specific purification

TAMRA Initial Final îeight Observed changes


weight weight loss
TAILA [oil] 995 g 995g No loss Tamra became soft and
blackish discoloration
TAKRA 993g 1.025kg Àain-32g Tamra had dark grey layers that
[buttermilk] peels easily
ÀOMUTRA 1.025kg 1.015kg Loss- 10g Àrayish black ,fragile,
[cow¶s urine]
ARANALA 1.015kg 950g Loss-65g Àrayish layers peels easily,
KULATHA 950g 925g Loss-25g Tamra became still more brittle
[decoction]
NIRÀUNDI 500g 575g Àain-75g Tamra powdered easily , saltish
RASA [juice of taste and smell
vitex nirgundo]
c
c
c
c
c

  
c
c

KAJJALI   45 6c


INÀREDIENTS:  purified]   c± 500g,  c 
-500g
Procedure:    c*c   and  
was done till the jet black powder was
obtained
Observation: triturating done for 62 hours , prepared kajjali was jet black lusterless and
soft
îeight- before trituration was 1000g
After trituration ± 970 g
Loss ± 30g

E] KAJJALI

c
c
c
 c c7;8c5 6
REFERENCE: "  c    c++
METHOD: incineration   
EQUIPMENTS: ’  c  [mortar pestle] ,       c[cowdung cakes]
INÀREDIENTS: ’   c± 2pts[300g],  c  [150g] ± 1pt ,lemon juice-qs
PROCEDUREDc
  and lemon juice paste was applied on copper and subjected to
incineration process by heating with 1000 cow dung cakes each time and after 10th 
genuine copper ash was obtained which passed all the classical   c 
  [tests for
genunity].

F] TAMRA BHASMA AT 10TH PUTA

6 
c
c

OBSERVATIONS:

 c First Second Third fourth Fifth Sixth seventh eight ninth Tenth
’   c 300g 250g 240g 240g 230g 226g 210g 196g 170gg 220g

c
 c 150g 125g 120g 119g 115g 113 105g 98g 85g 110g

c
, c 450g 375g 360g 360g 350g 115g 107g 102g 175g 220g
* c
 c
, c 145g 120g 119g 115g 113g 105g 98g 97g 107g 108g
*c
 c
(  1000 1000 1000 1000 1000 1000 1000 1000 1000 1000
 cc
# blac Blackish Brown Shine Black Black Browni Black Brown
 c kish with ish reduce and ish sh on color tinge
gree green and black d easily brow powder ed Black
n brown and compa powd n ing chakr ish
tinge soft rativel ered ikas brow
than y n
before Chakri
kas
soft
and
easily
breaka
ble
  c - - - (   (   (   (      P   Pc

   c      &c c   c c
 cc )c .c & 1+0c   c c
c*c "
 /0c c c  c 
    c c c c
c*c  c c c c
  c -c c -c
c

M 
c
c

   
c cc

After the sample was prepared the antibacterial study was conducted in MICROBIOLOÀY
DEPARTMENT OF JSS MEDICAL COLLEÀE MYSOREcc


 4ccc

PREPARATORY PROCEDURES:

A] Four different methods were adapted to make   c  suitable for this study.
1.c Sample I ± Preparation of stock solution by adding 1g of   c  sample in 10
ml of distilled water
2.c Sample II - Preparation of   c  paste by triturating well .
3.c Sample III - Pure filtrate was obtained from the original stock solution
4.c Sample IV - Solution obtained by diluting pure Filtrate with equal amount of distilled
water
B] Control groups :
Positive control group : In which bacterial suspension was maintained to check the
viability of organisms
Negative control group: were only distilled water for validation of the test was used .
C] Àentamycin in different solution with particular dose were used for comparison at
the same time.
D] A suspension was prepared with standard strains of E.coli and staphylococcus.
c

c cc4cc

 c c cc
 c c7:<8c

The Antibacterial action of the preparedc  c  cwas tested against both gram positive
cocci and gram negative bacilli . MIC & MBD of the sample was determined by following
NCCLS guidelines.

In the present study ATCC strains of gram positive bacteria (Staphylococcus Aureus) and
gram negative Bacilli (E coli) was selected as these are the common agents of infections
including Nosocomial infections.

Xc Sample I [À] -  c   was made into fine powder and stock solution was
prepared by adding 10ml of distilled water to 1g of sample.
From this stock solution different dilutions of   in decreasing concentration
was prepared ranging from 0.1g to 0.0125g /ml of nutrient broth in this way two sets
of test tubes was prepared and named as set A and set B

To each of these set of test tubes 1 loop full of each bacterial suspension [staph and E.coli]
matching to 0.5 mcfarland turbidity standard was added. set A test tubes were inoculated
with ecoli and setB was inoculated with staphylococcus . All the test tubes were then
incubated at 37 c for 18-24 hrs.

 
c
c

À] BROTH DILUTION SAMPLE I H] BROTH DILUTION SAMPLE II

Sample II [F]-    paste was prepared by triturating well with known quantity of
distilled water and dilutions were prepared with decreased concentrations of sample. Two
such sets of test tubes were sample concentration ranging from 5mg to 0.15mg/ml of nutrient
broth were made and named as set a and set b. these sets were also inoculated with bacterial
suspensions in the same manner and incubated at 37c for 18-24 hours.

Similarly the III sample - pure filtrate and the IVsample ± dilute solution of filtrate were
prepared and 1ml of each sample was taken with one ml of nutrient broth in different test
tubes and each of these were inoculated with bacterial suspensions and incubated for 18 hrs.

Negative and positive control groups and gentamycin dilutions were also prepared at the
same time .

After 18 hrs of incubation test tubes were taken out and wet mount preparation was done for
each of the dilutions and examined microscopically for viability of the bacteria. At the same
time from each test tubes and also from the control group one loop full of material were
aseptically inoculated on a blood agar plate. And all the blood agar plates were incubated at
37c for 18 hours.
On the third day blood agar plates were examined for presence or absence of bacterial growth
[colony formation] which shows no action or effective action of the samples respectively.
c
  cc4 After 18 hours of incubation of blood agar plates.
Blood agar plates ECOLI(Àram -ve bacilli) STAPHYLOCOCCUS(Àram +ve
cocci)
Positive control group Àrowth present Àrowth present
Negative control group Àrowth absent Àrowth absent
Àentamycin dilution test Àrowth absent Àrowth absent
tubes
Sample I [0.1g ± Àrowth absent [I] Àrowth absent [I]
0.0125g]
Sample II [5mg ± Àrowth absent in 5mg Àrowth absent in
0.15mg] 2.5mg [J] 5mg,2.5mg,1.25mg [K]
Sample III [pure filtrate Àrowth absent [L] Àrowth absent [L]
of sample]
Sample IV [ dilute filtrate Àrowth absent Àrowth absent
solution]
ë 
c
c

c
c
c
c
c
c
c
c
cccccccccccccccccccccccc
cc
5 6cccccccccccccccccccccccc ccccccccccccccccccccccccccccccccccccccccccccccccccccccc 56c
c
c
c
c
c
c
c
c
c
ccccccccccccccccccccccccc c
56ccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccccc5
6 c
c
c
 
4c
c
Xc The pure bacterial suspension or the positive control group showed bacterial growth.
Xc The negative control group containing only distilled water showed no bacterial growth.
Xc The gentamycin showed MBD[minimum bactericidal dose] at 0.01mg/ml
concentration
Xc The   c   c solution inhibited bacterial growth at 2.5mg/ml concentration for
Ecoli and
growth of staphylococcus was inhibited at 1.25mg/ml concentration.
Xc The   c  cfiltrate inhibited the bacterial growth in equal concentration even
after dilution.

  4cc
Xc Antibacterial action of  c  c 'c
  c
   c   properties
as said in ancient literature is established
Xc  c  cwas used in treatingc
 c
     c and 
 c
  .
Xc Previous studies onc  c  has proved its hepato protective ,antioxidant and
antiulcerogenic effect.

    
c
c

Xc  c  cgiven in appropriate doses would have very minimal or no side effects.
Xc In the present study its anti bacterial action on Ecoli and staphylococcus was made .
Xc By the Observations of the results it is proved that   c  chas antibacterial
action and has bactericidal effect on both the bacteria.
c

 4cc
Xc  c  is mentioned in treatment of 
c c
 c
  and 
 

 '
Xc Previous studies on   has shown that   is hepatoprotective in nature and
when prepared classically its toxic effects can be nullified.
Xc Its internal administration with an appropriate dose it may have less or no side
effects.
Xc Here the studies shows that   c   [4 samples] has a good amount of
antibacterial effect .
Xc Further research is required to find its exact therapeutic dose and with clinical studies
on infectious disorders it may be used as an ideal medicine in certain bacterial
infections.

c
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c

   :
(1)c Àopalkrishnabhatta-Rasendra Sara Sangraha, Dr. Ashok Satpute Edition 1st 2003
Choukambha Krishnadasa Acedamy Varnasi Chapter 1 Page194
(2)c Vagbhata, Rasaratna samuchachaya, edited by Kaviraja Ambikadatta shastri 8th ed
Varaanasi Choukmba samskrita samsthana:1988,5th chapter shloka 3/23
(3)c Panditha Narahari of Raja Nigantu Edited by Dr. Indradeo Tripathi 3rd Edn.
2003.Varanasi Choukambha Krishnadas Academy Suvarnadivarga sloka-18 Page-432.
(4)c Sadananda Sharma, Rasatarangini edited by Kashinath Shastri 11th ed. New Delhi:
Motilala Banarasidas Publication; 2004 17th Taranga Sloka-1 Page No-408.
(5)c Acharya Vishwanatha Dvivedi Bharatiya Rasashastra Edition 2nd , 1987, Shri, Sharma
Ayurveda Mandira Datia Jhansi Varanasi Nagpur.
(6)c Budheva mukharji Rasajala nidhi vol 2, 4th edition, Varanasi; Chawkhambha
Publishers;2004Chapter ± 4th Tamra prakarna page No-273.
(7)c Acharya Madhava- Ayurveda Prakasha, Sri. Àulrajsharma Mishra.Edn 2nd reprint 1999,
Choukambha Bharti Academy Varnasi Chap3. Sloka- 112Page 367
(8)c Dhanvatari Nighantuh Edited by Prof. P.V.Sharma Translated by Dr.Àuruprasada
Sharma 3rd edtion 2002 Varanasi Chokhambha Orientalia Publication. Shasta varga
sloka-9 page No-180.
(9)c Yadavji Trikamji Achary-Rasamritam. Dr. Damodhar Joshi Edition-1st 1998
Choukambha Sanskrit Sansthan Varanasi Chap 3 Shloka -34 Page 43.
(10)c Bhavamishra Bhavaprakash Nihgantu Edited by Dr. À.S.Pandey 9th edition,
Varanasi; Chaukhambha Bharathi Academy; 1993 Dhatwadi varga sloka23 page No 6
(11)c Dr.K.M Nadakrani ± Indian Materia Medica Volume II ± A.K Nadakarni Edition 2nd
Reprint 1992 Popular Prakashana Bombay ,Page 47
(12)c VaidyaVasudeva Mulashankar Diwedi- Parada vijnanam, Edition-2nd 1978.Sri
Sharma Ayurveda mandira Varanasi Chap1.
(13)c Minerals of India by Meher. D. N. îadia
(14)c Field Cruied to Rocks & Minerals of the world by Larousse
(15)c Dr.K.M Nadakrani ± Indian Materia Medica Volume II ± A.K Nadakarni Edition 2nd
Reprint 1992 Popular Prakashana Bombay ,Page 48
(16)c Vagbhata, Rasaratna samuchachaya, edited by Kaviraja Ambikadatta shastri 8th ed
Varaanasi Choukmba samskrita samsthana:1988,3rd chapter shloka 23, p.100
(17)c B. N. Àhosh -Pharmacology, Materia Medica and Therapeutics , Edi.
XVIIIth Hilton & Co.

ÿ    
c
c

(18)c Pharmacopoeial Standards for Ayurvedic Formulations, Revised Edi., 1987,


CCRAS, New Delhi
(19)c Satoskar R.S. and Bhandarkar S.D.,Pharmacology Pharmacotherapeutics Edi.
XIIth, 1991, Popular Prakashan, Bombay
(20)c R. Ananthanarayanan, C. K. Textbook of Microbiology Edi. IVth 1990 by Jayaram
Paniker, Orient Longman Limited, Madras
(21)c Clayton Thomas-Taber's Cyclopedic Medical Dictionary, Edi. XVIIIth, 1998,
Jaypee Brothers Medical Publishers, New Delhi.
(22)c Norman Evers and À. D. Elsdon. C. -The analysis of drugs and chemicals Company
Ltd., 1929

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