Standardization of process for increased production of pure and potent tetanus toxin

Abstract

Objectives: The aim of the study was to increase the yield of tetanus toxin in short time fermentor cultivation and also to produce pure and potent tetanus toxin replacing initial nitrogen source (N.Z Case) with papain digest broth in the modified Mueller Miller medium (MMMM). Materials and Methods: A fermentor, using a vibromixer and optimum supply of sterile air to the headspace of the fermentor to flush out the accumulated gases was used. The MMMM containing initial N.Z Case was replaced with papain digest broth was used successfully. Results: It was found that under optimal conditions of temperature, vibromixing, surface aeration, and an alkaline pH favored toxin release. Furthermore, to enhance the production volume, fermentor culture is more suitable. The tetanus toxin was produced with good Limes flocculation (Lf) titre and high antigenic purity. A significant increase in the tetanus toxin yield in short time cultivation (about 5 to 6 days against 8 days) was noticed even with MMMM containing papain digest broth instead of N.Z.Case. Conclusion: For large-scale production of puri¬fied and potent (antigenic purity) tetanus toxin, the use of fermentor technology can be utilized under optimal conditions. The production medium using indigenously available ingredients containing high level of aminonitrogen as in the case of PDM can be substi¬tuted in place of N.Z Case, which is being imported and expensive, in addition to lot-lot variation.

Keywords

• Clostridium tetani, modified mueller miller medium, papain digest, limes flocculation

Standardization of process for increased production of pure and potent tetanus toxin

Öz

Amaç: Bu çalışmanın amacı nitrojen kaynağı papain digest broth içeren N. Z Case yerine modifiye Mueller Miller medium (MMMM) kullanarak tetanoz toksin üretiminin artırılabileceğini göstermektir.Yöntemler: Clostridium tetani’nin sabit pot kültürü metodu bir vibromikser ve fermentör boşluğuna steril hava sağlayan düzenek kullanılarak fermentörde kültüre edilmesi esasına dayanan daldırma kültürü metodu ile değiştirildi.Bulgular: Isı, vibromiksing, yüzey havalandırması ve alkalin pH’nın optimmal şartlarının sağlanması ile toksin salınımı sağlandı. Ek olarak üreme hacminin arttırılmasında fermentör kültürü daha uygun idi. Üretilen tetanoz toksininin Limes flokülasyon (Lf) titresi iyi ve yüksek antjienik saflığa sahip idi. Tetanoz toksin üretiminde N. Z Case yerine modifiye Mueller Miller medium (MMMM) kullanarak kısa süreli kültivasyonla (8 güne karşılık 5-6 gün) belirgin artış sağlandı.Sonuç: Büyük miktarlarda saf ve potent (saf antijenik) tetano toksin üretimi için optimal şartlarda fermentor teknolojisi kullanılabilir. Saf ve potent tetanoz toksini eldesi için N.Z Case yerine papain dijest broth besiyerinin kullanılabileceğini düşünmekteyiz

Anahtar Kelimeler

• Clostridium tetani, modifiye mueller miller besiyeri, papain dijest, limes flokülasyon

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