Pharmaceutical Microbiology

Sterilization

Sterilization refers to any process that removes, kills or inactivates all forms of life and biological agents present in or on a specific surface, objects or fluid. Sterilization achieved through various means including heat, chemicals, irradiation, high pressure and filtration.

Sterilization Technique

1. Physical Method of Sterilization:

⇨ Moist heat sterilization

⇨ Dry heat sterilization

⇨ Radiation sterilization

⇨ UV-rays sterilization

⇨ Ionization sterilization

Moist heat sterilization

⇨ Moist heat sterilization is done using Autoclave (Steam under pressure)

⇨ Mode of Action: Denaturation or coagulation of protein

⇨ Satutated stream under pressure

Most condition: 121°C /15 Ibs-psi/ 15-20 min

⇨ 115-120°C/ 30 min

⇨ 121-124°C/ 15 min

⇨ 126-129°C/ 10 min

⇨ 134-138°C/ 5 min

⇨ For clostridium tetani: 115°C at 30 min

Dry heat sterilization/ Depyrogenation

Instruments: Hot air oven, Incenarator, Flaming, Glass bead sterilization

Mode of Action: Oxidative free radical damage of microorganism/ Destroy the ability of microorganism

Temperature & Time conditions fir dry heat sterilization:

⇨ 160° at 2 hours

⇨ 170°C at 1 hour

⇨ 190°C at 6-12 min

⇨160-180°C at 2-3 hours

* Depyrogenation seeks to destroy the chemical activity of the by-products, pyrogens or endotoxins.

Pyrogen 

Pyrogens are the metabolic products of microorganism that are produced by all microorganism like gram-positive,  Gram-negative bacteria and fungi.

⇨ Pyrogens are bacterial cell wall fragments. They are not bacteria. Typicall they are complex carbohydrates.

⇨ Pyrogen is a fever inducing substance.

⇨ Pyrogens can be classified into two groups: endotoxins and non-endotoxin pyrogens. Enfotoxins are substances found in gram negative bacteria while non-endotoxins are found in gram-poditive bacteria.

Pyrogen Test

The pyrogen test measures the pyrogenic load in parenteral products. It is a regulatory requirement for the release of sterile parenteral products.

Types of Pyrogen test:

⇨ Rabbit test

⇨ LAL test (Limulus amoebocyte Lysate test)

LAL test

It is an in vitro test for bacterial endotoxin or pyrogens use a lysate of amoebocyte from the horseshoe crab limulus polyphemus.

Endotoxins

Endotoxins are the main component of outer membrane of the cell wall of gram-negative bacteria. The main function of endotoxins in bacteria is structural and protective action.

Pyrexia

Pyrexia or fever in human body is a body temperature above the normal range due to an increase in the body temperature set point in the hypothalamus.