Factors affecting microbial spoilage of

pharmaceutical products
1. Type and size of contaminant inoculum

2. Nutritional factors: Most acute pathogenic

bacteria have specific nutritional requirement
so they are unlikely to grow in pharmaceutical
preparations. Some have minimal nutritional
requirements e.g. Pseudomonas can grow
significantly in deionized water.
3. Water activity (Aw): By decreasing the water
activity (the readily accessible water) in the
product, we can prevent microbial growth. Aw
can be lowered by increasing the solute
concentration in the pharmaceutical product
e.g. sucrose, sorbitol, fructose.

4. Storage temperature: Reconstituted

suspension or syrups and multi-dose eye
drops are usually instructed to be kept in the
fridge (4-8°C) after first use to prevent
microbial growth. Water for Injection should be
kept at 80°C after distillation in the
pharmaceutical plant and before packing to
avoid contamination.
5- Redox potential (=Oxidation reduction
balance) of the pharmaceutical product: It
is due to the presence of oxidizing and
reducing agents. It affects the ability of
microorganism to grow in the
pharmaceutical product.

6- pH: Above pH 8, spoilage is rare. Yeasts

and mould can survive in pH 3-4. Very low
pH is not usually used in medicines.
7- Packaging design: Self-
sealing rubber wads in
multidose injection containers
to avoid contamination after
using needles.

•Narrow nozzle,
collapsible screw-
capped tubes are
preferred over wide-
mouthed cream jars.
 The ideal pharmaceutical
preparation should be:

1. Effective
2. Safe
3. Stable during shelf life
4. Elegant
Quality Assurance (QA) and Good
Manufacturing Practice (GMP)
• Quality Assurance (QA): It refers to the
whole arrangements made to ensure that
the final product is of the quality required
for its intended purpose. It consists of
Good Manufacturing Practice (GMP) plus
factors such as original product design
and development (R & D) and post-
marketing surveillance.
Good Manufacturing Practice (GMP)
or current Good Manufacturing Practice (cGMP)
or Good Pharmaceutical Manufacturing Practice
(GPMP):

It is the part of Quality Assurance (QA) that aims at

ensuring the product is consistently manufactured to a
quality appropriate for its intended use. It includes the
control of ingredients, plant construction, training of
personnel, process and equipment validation,
production and cleaning.
• Quality Control (QC):
It is the part of GMP that ensures that at each stage of
manufacture the necessary tests are conducted and
documented and the product is not released until it
passes these tests.
e.g. Test for endotoxins in sterile pharmaceutical
products

In-process Control:
It refers to any test on the product, the environment
and the equipment that is made during the
manufacturing process. e.g. testing that an
autoclave is functioning correctly.
 Quality Assurance (QA)

Good Manufacturing Practice

(GMP)

In-process Quality Control

Control (QC)
• Before a product is released to the
market it must pass the Quality Control
tests:
1. For sterile pharmaceutical products:
a) Sterility testing
b) Endotoxin or pyrogen testing.

2. For non-sterile products: results of the

Microbial Limit Test (MLT) must be
acceptable according to pharmacopeial
guidelines.
 Contamination of a pharmaceutical
product after manufacturing
• This happens for sterile multi-dose
products (injection and eye drops)
• It also occurs in non-sterile uni-
dose and more frequently in non-
sterile multi-dose preparations.

• Sources of contamination include:

1. From the patient: e.g. saliva, nasal
discharges, skin.
2. From the air.
Some ways to reduce microbial contamination
after manufacturing and during use:
1. Dispensing uni-dose preparation whenever
possible. (e.g. tablets in strips not in a jar,
powder in sachets not in bulk, uni-dose eye
drops).

2. For ointments and

creams: Use of collapsible
tubes rather than wide-
mouthed jars. Metal
collapsible tubes is better
than plastic ones as they
don’t pull air after
collapsing.
3- Incorporation of preservatives:
Preservatives are added to multi-dose
preparations to prevent or limit any microbial
contamination that may happen during normal
storage and use of the product. Adding
preservatives must not be a substitute for GMP.
Examples of preservatives:
a) Acids and esters: e.g. Benzoic acid, sorbic acid.
The active moiety is the undissociated acid so
they are active in acidic pH below pH 5.

HA H+ + A-
(Undissociated acid)
Esters of p-hydroxybenzoic acid (Parabens):
These are methyl, ethyl, propyl or butyl esters.
Usually used in combination for emulsions
where methyl ester protects the aqueous phase
and propyl and butyl esters protect the oily
phase. They have antimicrobial activity in acidic
and neutral pH. They have broad anti-fungal
activity but less active against bacteria esp.
Pseudomonas which can utilize Parabens as a
carbon source.
b) Quaternary ammonium compounds (QACs):
e.g. Benzalkonium chloride, cetrimide,
cetylpyridinium chloride. They are active at
neutral or slightly alkaline pH. They are
completely inactive at pH below 3.5.
They are active mainly against Gram positive than
Gram negative bacteria. They have limited
antifungal activity mainly fungistatic rather than
fungicidal.
Testing the efficacy of antimicrobial
preservative
• The antimicrobial activity of the preservative is
tested in its final preparation.

• Preservative antimicrobial activity can be

affected (enhanced or diminished) by the active
ingredient of the preparation, by the formula,
and by the container and closure used.
 Challenge test (BP 2010)
• The test consists of challenging the preparation
with an inoculum (105 – 106 microorganism per
ml or per gram of the preparation) of a suitable
microorganism, then storing the inoculated
preparation at a prescribed temperature and
taking samples at intervals to count the
microorganism.

•The sampling period usually extends to

28 days after inoculation.
•The preservative would be adequate if
there is a significant fall or at least no
increase in the number of microorganism
Challenge test BP 2010 (cont.)

• Microrganisms used for the challenge test:

Standard strains of Ps. Aeruginosa, S. aureus,
Candida albicans and Aspergillus niger.

• Any other microorganism that is likely to

contaminate the preparation can be used in the
challenge test e.g. E. coli can be used for oral
preparations.

• Note that for testing the microbial count, it is

essential to remove the antimicrobial effect of
the preservative by dilution, membrane filteration
or adding inactivating agent (Quenching agent).
Challenge test BP 2010 (cont.)
Challenge test BP 2010 (cont.)
 “Capacity test” for preservatives:
• It is a challenge test where the product is
repeatedly challenged at intervals until the
preservative system fails.
• It simulates the situation of re-
contaminating a pharmaceutical product
by patients e.g. creams.
General notes:
• Areas in a pharmaceutical plant are
classified according to the air quality to
black (the worst), grey, white and aseptic
(sterile) areas.

• Areas with higher air quality are kept at a

positive pressure of air compared to
adjacent areas of lower air quality to
prevent any influx of contaminants from
low grade areas.