STERLIZATION

Sterilization of parenterals

Aseptic technique:- is the preparation of pharmaceutical products

from sterile ingredients by procedure that exclude the access of viable

microorganism into the products

It is used for those would be adversely affected by being subjected to

sterilization

Sterilization = complete removal or killing of all viable

microorganism and their spores

Microorganism can be

categorized into three types

Easy to kill by dry or moist heat

Susceptible to moist heat and resistant to dry

heat

Susceptible to dry heat and

resistant to moist heat

1- Dry- heat sterilization

2- Steam sterilization

3- Sterilization by filtration

4- Gas sterilization

5- Sterilization by ionization

• However the applied method for sterilization the product must pass the sterility test.

Important to:-

• As a proof of the effectiveness of :

• the applied method,

• Performance of the equipment

• Performance of personal

1- Dry sterilization (Oven method

• Less effective in killing M.O , • so higher temp and longer time of exposure are required. • Temp of 140 ° for 3 hrs or 260 ° for 45 mins are efficient to

kill spores and vegetable M.O

• (due to dehydration of microbial cell followed by a slow burning or oxidative process)

Dry sterilization use Ovens The modern ovens are supplied by heated, filtered air that is

uniformly distributed throughout the oven chamber

1- Used for heat stable non-aqueous preparations, powders, 2- small individual units to permit free circulation of heated air

3- equipment (glass , porcelain, not for plastic) 4- heat stable powders (kaolin, sulfur, Zno…) 5- Surgical instrument 6- Substances that can not sterilized by moist heat (Fixed oil, Glycerin, petrolatum, paraffin)

1- Dry heat method usually applied to:

2- Moist heat sterilization • Based on heating in saturated steam under pressure • More effective in killing M.O than dry method.

• The BP & EP recommended 121 ° C maintained throughout the load for 15 mins as the preferred combination for this method of terminal .

• So pressure is applied to achieve higher temp. Autoclaves that employ steam under pressure which ensure destroying

spores bacteria within 20 mins at 15 pounds (temp = 132)

a- Moist heat cause denaturation or coagulation of

bacterial cell protein at much lower temp

B-Thermal capacity of the steam is greater than that of

hot air

Steam sterilization is applicable to

1. Aqueous solution packed in sealed containers as ampoules

2.Bulk solutions

3.Glassware

4.Surgical dressing and instruments

5.Can be applied with temp lower than 100 °C (suitable for substances not withstand higher temp.

Not suitable for :

1.Oils, fats

2.Preparation that not penetrated by moist

3.Powders that may be damaged by moisture

3- Sterilization by filtration

• Used to remove particles including M.O from solutions from or gasses (air) by :

• - adsorption on the filter paper

• Sieving mechanism

• The applied filter paper should not remove the desired constituents or add any undesired constituents.

• Applied for heat sensitive solutions

• Filters are two basic type:

• Depth and membrane

Based on combination with

pathway and adsorption to

retain M.O or particles

Based on sieving

through the filter

pores

Membrane filter media Millipore filter

• Plastic filter

- Cellulose nitrate

- Cellulose acetate

- Acrylic copolymer with nylon

- Polysulfonate

Sintered glass or

metal filter

Used when durable

characteristics are

required

laboratory for filtration sterilization of medium components. The filter is

placed (aseptically) on the glass platform, then the funnel is clamped and the fluid is

drawn by vacuum into a previously sterilized flask. The recommended size filter that

will exclude the smallest bacterial cells is 0.22 micron.

Gas sterilization

• Applied for heat & moisture sensitive materials

Ethylene oxide Propylene oxide

Flammable gases usually applied after

mixing with inert gases

Sterilization time 4-16 hrs

Ethylene oxide (ETO) is the most commonly used form of chemical

sterilization.

Due to its low boiling point of 10.4ºC at atmospheric pressure, EtO)

behaves as a gas at room temperature.

The sterilization process is carried out in a specialized gas chamber..

An ethylene oxide sterilization gas chamber

• ethylene oxide interfere with the metabolism of bacterial cell .

mode of action:

Application:

Applied for thermolabile

materials, without damage

medical and surgical supplies

such as -- catheters, needles,

plastic disposable syringes in

their final packaging

The gas can diffuse

through packaging

materials and containers

made of paper, fabric, a

range of plastics & rubber

And diffuse out again after

sterilization

EtO chemically reacts with

amino acids, proteins, and DNA

to prevent microbial

reproduction.

Sterilization by ionizing radiation

•applied by electromagentic waves e.g – •gamma rays --- Infrared radiation --- X- rays •Ultraviolet light (wavelength near 265 nm are the most effective, as they are strongly absorbed by nucleoproteins) • (Disadv. Of U.V = poor penetrating power) •Applied to clean air , water in thin layers

Ionizing radiation by gamma radiation have high penetrating power and high sterilizing efficiency, little or no damage effect on irradiated materials

Is the most common in UK & used radioisotope coblet 60Co

But still limited technique bec: require of specialized equipment

undesirable effect on products and containers

Mode of action:- ionizing radiation cause excitation, ionization

and where water is present, free radical formation

Free radicals are powerful oxidizing agents&

capable of damaging essential molecules in living cells

WEEK 12

•VALIDATION OF

STERILITY

Validation of sterility

• Regardless the applied method for

sterilization, Sterility test must confirm the

absence of viable micro-organisms in samples

taken from production batch

1- Sterility test

Direct transfer

to test media

membrane filtration

technique

Direct transfer

to test media

Aseptically directly transfer a

specified volume of test

material

into culture media

Incubate for 14 days examine the media visually for growth (turbidity) at

least on

3rd, 4 th, 5 th and 8th

No microbial growth = sterile

membrane filtration

technique based on using aseptic membrane filter (pore size

not greater than 0.45 um) after filtration the

membrane filter is incubated aseptically into

appropriate media (proper temp. and time)

It is applicable for ---- aqueous solution,

--- soluble solids

---- and oils & oily solutions

Pyrogen testing

• Pyrogens = organic substances

• can removed by oxidizing them to easily eliminated gases or to nonvolatile solids

that can removed from water by fractional distillation.

Oxidizing agents = potassium permanganate + small amount

of barium hydroxide

( impart alkalinity + PPt barium salts of any acidic compd)

Test for Pyrogen The BP pyrogen test involves measuring the rise in temperature of healthy mature rabbits. The temp. is recorded after the rabbits have been IV injected with sterile solution of the test subs. 1- tools (glassware, needles, syringes free from Pyrogen) heated to 250 °C to be free from pyrogen 2- exclude any rabbit with unusual response 3- Warm product to 37 °C 4- inject into ear vein of 3 rabbits 10 ml/kg body weight) 5- record temp at 30 mins (from 90 mins before the test and to 3 hrs after injection )

If no rabbit shows any rise in temp (0.5 or more) =

product free from pyrogen

If any rabbit shows any rise in temp (0.5 or more) =

continue the test using five other rabbits

If not more than 3 rabbits show rise in temp (0.5 or more) =

or sum of 8 rabbits maximum temp = 3.3 C

the material under examination meets the

requirements for the absences of pyrogen

Parenteral quality control tests

1. Drug content and volume in container

2. Sterility test

3. Pyrogen test

4. Leaker testing and sealing verification

5. Clarity testing and particulate analysis