B. Amsden CHEE 440

STERILIZATION

The complete destruction of all living organisms and their spores or their complete removal from the formulation.

All parenterals, as well as otic, nasal, ophthalmic solutions, must be sterile,

includes packaging materials

methods steam dry-heat filtration gas ionizing radiation

Viruses

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Bacteria

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Bacterial Endospores

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Yeast and Mould

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aspergillus penicillium

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Sterilization Methods

steam sterilization conducted in an autoclave method of choice 121 C, 20 minutes

dry heat less effective than moist heat 170 C, > 2 hours used to sterilize equipment

B. Amsden CHEE 440

Heat Resistances

method determined by nature of micro-organism and formulation ingredients

protocols are developed based on reference organisms of known sterilization resistance

micro-organisms possess different resistances to heatcell type resistance

bacterial endospores may survive hours inboiling water

fungal spores death at > 80 C in 60min.

yeasts survive 20 min at 60C

bacteria D60 = 1-5 min

viruses

protozoa, algae

rarely survive > 30 minat 60 Crapidly die at T > 40 C

B. Amsden CHEE 440

Steam Sterilization

Determination of steam sterilization protocol first select a reference organism determine its D value under defined conditions establish initial bioburden calculate time of exposure at 121 ºC

B. Amsden CHEE 440

Sterilization Methods

Filtration used for heat sensitive solutions pore sizes range from 14 to 0.025 m

Gas sterilization used in hospitals and industry for thermolabile materials ethylene oxide, propylene oxide used 500-1000 mg/L, 4 - 16 hours at 55 ºC, 50-60 %RH need to remove absorbed gas after sterilization

Ionization use of gamma rays limited application

B. Amsden CHEE 440

Kinetics of Cell Inactivation

death rate follows first order kinetics

N t =N0e−kt

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D value

• decimal reduction time• time required for 90% reduction in

cell count

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Z value

degrees of temperature/dose change required to achieve a tenfold decrease in D

B. Amsden CHEE 440

Example

Using G. stearothermophilus as a reference organism (D121 = 1.5 min), and an initial bioburden of 1000 spores, determine the sterilization time required to achieve a 1 in 1,000,000 chance of a spore surviving using a steam autoclave.

B. Amsden CHEE 440

De-pyrogenation

Emphasis is placed on prevention of introduction of pyrogens

Removal glassware, metals

• 250 ºC for 45 minutes water

• oxidation to nonvolatile solvents using potassium permanganate, then distill

• reverse osmosis plastics

• protect against contamination