1. QUALITY CONTROL OF SNAKE ANTIVENOM
DR.M.V.KHADILKAR
Technical Director,
Premium Serums & Vaccines Pvt.Ltd.,
Narayangaon,Dist.-Pune
Maharashtra

2. PRODUCTION PROCESS Equine(Horse/Mules/Ponies) procurement
Quarantine/Complete Health screening
Regular periodic Immunization with snake venoms
Periodic Titer Testing
Plasma collection & storage
Plasma Fractionation
Batch pooling
Sterile Filtration
Containerization & Lyophilization
Labeling /Inspection
Finished Product Testing
Marketing

3. Antivenom quality and safety depends on
1.The control of source materials 2. Manufacturing procedures

4. Basic Raw Materials in Antivenom Production
Snake Venoms
Hyperimmune Plasma

5. Objectives of the purification process
High safety and efficacy
No impairment of the neutralizing activity during purification
Maintenance of high purity
Free of microbial contaminants and endotoxins
WHO Workshop,

6. precautions in Venom production
Venom is to be obtained from snakes in a controlled and documented manner. It should be properly preserved and tested for quality Venom being the prime raw material for the production of Antivenom ,it is a key factor in the production of effective antivenom.

7. Monitoring of immune response Proper collection process
Plasma production
Animal health
Monitoring of immune response
Proper collection process
are the greatest contributors to good quality plasma production

8. Collection of plasma
Maintenance of healthy & disease free herd of animals
Cleanliness of premises and complete asepsis during procedure.
Use of sterilized collection sets & containers under controlled environmental conditions
Properly designed clean room to separate plasma and operations carried out by trained personnel.
Storage in proper temperature controlled rooms.
Plasma collection vessels must be labeled with date, plasma volume, horse numbers and anti-coagulant use details.
AVOIDS BIOBURDEN-HELPS IN CONTROLLING ENDOTOXINS

9. ENZYMATIC CLEAVAGE OF IMMUNOGLOBULIN MOLECULE

10. COMMON METHOD USED FOR PLASMA FRACTIONATION in India
Pepsin Digestion to get F(ab’)2 fragments from whole IgG.
Precipitation of unwanted proteins by either Ammonium Sulfate or Caprylic Acid
Concentration & Diafiltration of F(ab’)2
OTHER METHODS
Use of whole IgG molecule
Papain Digested Fab fragments
Chromatography (Ion-Exchange/Affinity)

11. PLASMA FRACTIONATION
Use of closed system with no exposure to environment.
Least product hold time during process
Process validation-digestion, filtration, ultrafiltration
Strict supervision of critical process parameters such as Temp,pH,duration etc
Checks about cleaning & sterilization status
IPQC checks -

12. Quality Control TESTS DESCRIPTION pH PROTEIN PRESERVATIVE CONTENT
POTENCY
ABNORMAL TOXICITY
STERILITY
ENDOTOXIN
(SOLUBILITY & MOISTURE CONTENT)

13. ANTIVENOM SAFETY
Depends on 1. Type of antivenom -Whole IgG/ F(ab)2 / Fab Each fragment has different pharmacokinetics Efficacy reports not supported by clinical data 2. Dose 3. Route & Speed of administration 4. Manufacturing practices Hypersensitivity skin test has limited predictability value.

14. Reasons for reactions Contamination of plasma
Enzymatic digestion & removal of unwanted proteins
Formulation
Sterility
No data from Indian clinical trials is available, however , a lot has been reported by importing countries
Thought to vary from 3-80% with about 5-10% severe reactions
Reports are often subjective

15. EFFICACY MONOVALENT / POLYVALENT –Has its own advantages/disadvantages
LEVEL OF REFINEMENT-Enzyme digestion,Ion exchange chromatography/Affinity chromatography. Effect on cost????
VENOM
QUALITY
GEOGRAPHICAL ANTIGENIC VARIATION
SPECIFICITY/PARASPECIFICITY
CLINICAL MANAGEMENT-OUTCOME DEPENDS ON
EARLY START OF TREATMENT
ADEQUATE DOSES
SUPPORT SYSTEMS

16. SNAKE ANTIVENOM I.P. Name of Product : Snake Antivenom I.P.
(Polyvalent, Lyophilized/liquid, Equine origin Enzyme Refined Immunoglobulin)
Description of Product :Lyophilized 20.0 ml Vial
Along with 10.0 ml W.F.I. I.P. OR liquid 10 ml vial
Potency : When Reconstituted With 10.0 ml
of Sterile Water For Injection I.P.
1.0 ml of reconstituted mixture Neutralizes
0.60 mg. of Cobra Venom
0.45 mg. of Krait venom
0.60 mg. of Russell’s viper venom
0.45 mg. of Saw scaled viper venom
Nature and concentration
of preservative : Phenol /Cresol not more than < 0.25%
Storage conditions :Store in cool dark place, avoid
exposure to sunlight for lyophilized and to store between 2-8˚C for liquid product.

17. How much antivenom is needed
How much antivenom is needed?? ( Tumbare & Khadilkar,Journal of Bombay Veterinary college:2004,12(1&2):9-11)
Snake
Average Venom Yield in ml
(Liquid)
Average Venom Yield in mg (Lyophilized)
Theoretical dose of antivenom required for treatment
(10 ml vial)
Indian Cobra ml
(0.435±0.01) mg
(126.32±4.31) 21
Russell’s Viper
ml (0.412±0.02) mg
(75.98±3.66) 13
Indian Common Krait ml
(0.101±0.01) mg
(8.35±0.64) 2

18. INDIAN SCENARIO
It is estimated that, India currently produces about million Snake Antivenom vials per year
Almost 75-80% of total Snake Antivenom produced is supplied to different government institutions via tender procedure.
Current capacities can supply antivenom that meets about 60-70% of the requirements leading to unfortunate incidents of deaths
Many times the problem is compounded by improper distribution
This results in big demand-supply gap for this life saving drug

19. INDIAN SCENARIO
Many Indian companies have been regularly supplying this life saving medicine not only to its neighboring countries but also to countries in Africa and South Asia.
Due to the enforcement of various regulations after 2000, manufacturing of equine biological products was significantly reduced resulting in severe shortage of these life saving products.
Stringent regulatory requirements have also pushed up the production costs

20. INDIAN SCENARIO
India still has very low price as compared to many other countries
It has been observed that 2010 prices for a 10 ml vial of Indian polyvalent AVS range from about INR 300 to 500 (US$ 6.50–11.00), which is a fraction of the cost of a vial of CroFab antivenom in the USA (at over US$ 1900 per vial) or CSL antivenom in Australia (at US$ 1500 per vial)
AVS is supplied by Indian antivenom producers to government hospitals at Rs 115 per vial (US$ 2.50)
(Romulus Whitaker & Samir Whitaker in Current science, vol. 103, no. 6, 25 september 2012)
Most of the ASVS tenders were awarded at Rs Rs. 210 per vial in 2011

21. My views
Dose finding studies can give guidelines for treatment protocol based on syndromic approach.
One protocol for big country like India is difficult considering regional variation in venom composition.
Use of polyvalent antivenom is more practical and economical especially considering procurement and distribution system followed.
More legal sources of venoms should be established/promoted in different regions for its availability in production process.
The Indian antivenoms are reasonably priced.
There is limit to purification considering its direct effect on price.
Collaboration between Industry & Academic institutions is needed to improve yields & reducing costs.

22. THANK YOU