Ipqc tests for parentrals

IPQC TESTS FOR
PARENTRALS
MOSES.M
M.PHARMACY
256212886050
GUIDED BY: Dr. NIRMALA
Dept. of Pharmaceutics
Presented By:

Introduction to Parentrals:
 What are parentrals?
 Greek para = "beside" and enteron =
"intestine", because it bypasses the
intestines.

 Parentrals are the dosage form for conveying drug
by means of injection through the skin or mucous
membrane.
Parenteral drugs are administered directly into the
veins, muscles or under the skin, or more specialized
tissues such as the spinal cord.
Circumvents the highly efficient first line body defense
that is skin and mucous membrane.
Thus they should be free from microbial contamination
and should have high purity.

Definition:
 Parenterals are
◦ Sterile
◦ Pyrogen-free &
◦ Free from particulate matter
which are injected into the internal body
compartment.

Quality Assurance:
 It is the sum total of the organized
arrangements with the objective of
ensuring that the products will be of
the quality required for their intended
use.

GMP:
 Is that part of Quality Assurance
aimed at ensuring that the products
are consistently manufactured to a
quality appropriate to their intended
use.

Quality Control:
 Is that part of GMP concerned with
sampling, specification & testing,
documentation & release procedures
which ensure that the necessary &
relevant tests are performed & the
product is released for use only after
a.scertaining it’s quality.

In Process Quality Control
 IPQC means controlling the
procedures involved in manufacturing
of the dosage forms starting from raw
material purchase to dispatch of the
quality product in ideal packaging.
 It monitors all the features of the
product that may affect its quality and
prevent errors during processing.
 It is the activity performed between QA
and QC.

Importance:
 To minimize human errors.
 Provides accurate, specific and definite
description of the procedure to be
employed.
 It is a planned system to identify
materials, equipments processed and
operations.
 Is to detect the errors if and when it does
occurs.
 Is to enforce the flow of manufacturing
and packing operations according to
established routes and practice.

IPQC For Parenterals:
 Each filled vials and ampoules are
subjected to inspection for particles,
volume, cap or seal conditions.
 Assay: Drug content
 Clarity test: For particulate matter by
Visual method
Coulter counter method
Filtration method/Microscopic count
method
Light blockage method

Continued..
 Conductivity test
 Fill volume
 Leak test:
 Dye bath test
 Liquid loss test
 High voltage test
 Spark test

Continued..
 Sterility test:
Membrane filtration method
Direct inoculation method
 Pyrogen test:
Testing on Rabbits
Limulus Amoebocyte Lysate (LAL) test
 pH
 Label check
 Pass ability of Parenteral preparation
through needle(Viscosity)

Drug Content:
 Assay:
 Assay is performed according to the
method given in the monograph of that
parenteral preparation in the
pharmacopoeia.
 Assay is done to check the quantity of
medicament present in the parenteral
preparation.

Clarity test:
 VISUAL INSPECTION BY NAKED
EYE:
◦ Each injectable is inspected visually
against White and Black Backgrounds.
◦ The White backgroud aids in detection
of dark colored particles.
◦ The light reflective particles will appear
against the Black background.

Coulter Counter method
 The sample solution is added to an
electrolyte solution which is drawn
through a small orifice.
 As particle passes through the orifice it
displaces its own volume of electrolyte.
 Particle detected by the increase in
electrical resistance.
 Voltage pulses are proportional to the
particle size.
 Particles below 0.2μm can also be
detected.

Limits for detection of subvisible Particulate matter as prescribed in USP
Particle size SVP LVP
≥10μm
3000/containe
r
12/ml
≥25μm 300/container 2/ml

Microscopic count method
 Membrane filter and microscopes are
used.

Light blockage/Obstruction
method
 This method uses an electronic counter that
produces a light beam of high intensity.
 The solution is allowed to pass under this
bright light.
 A shadow is formed if a particle is present.
 The particles are counted by the no. of
shadows.
Limits for subvisible particulate matter as prescribed in USP
Particle size SVP LVP
≥10μm
6000/containe
r
25/ml
≥20μm 600/container 3/ml

Leakage Test
 Ampoules are subjected to this test.
 Leakage test not done for vials and
bottles.
◦ 1. using methylene blue solution/Dye bath
test
◦ 2. spark test

Leakage test (with methylene blue
solution)
 The ampoules are immersed in vacuum
chamber consisting of 1% methylene
blue solution
 A vacuum of about 27” (inch) Hg is
created for about 15 to 30 min
 This causes the solution to enter the
ampoules with defective sealing.
 The vacuum is released and ampoules
are observed.
 If a leakage is present, the solution in the
ampoules appear blue color.

Spark test
 The machine uses high precision
electrodes to inspect the full
circumference of the containers,
including the closure zone.
 All containers are presented inividually to
the electrodes.
 Any moisture that has penetrated
through capillary forces in a crack,
pinhole or just weak glass is registered
as a change in resistance.
 All products with a measured voltage
higher than a defined maximum value
are separated from the good products.

Advantages of HVLD test
 Vials and ampoules also can be
tested.
 High accuracy of inspection.
 High speed of processing.

Sterility Test
 Sterility testing attempts to reveal the
presence or absence of viable micro-organism
in a sample number of
containers taken from batch of
product. Based on results obtained
from testing the sample a decision is
made as to the sterility of the batch.
 Sterility testing is made after the
product exposition to the one of the
possible sterilization procedures.

Principle
 Sterility test is based on the principle
that when micro-organisms are
supplied with nutrient medium and
water, and incubated at favorable
temperatures, they multiply.
 The presence of micro organisms can
be identified by turbidity in the clear
medium.

Culture Media
 The culture media used for sterility
test must be capable of promoting the
growth of a wide range of micro
organisms.
 Types of media-
◦ 1. Fluid thioglycollate medium (for
anaerobic bacteria).
◦ 2. Soyabean-casein digest medium (for
aerobic bacteria and fungi).
 Incubation of media: 14 days at 30-
35oC

Fluid Thioglycolate Medium
Ingredients Quantity for 1000 ml
L-cysteine 0.5 g
Sodium chloride 2.5 g
Dextrose 5.5 g
Agar 0.75 g
Yeast extract 5.0 g
Pancreatic digest of casein 15.0 g
Sodium thioglycollate 0.5 g
Resazurin (0.1% fresh
1.0 ml
solution)
Distilled water Upto 1000 ml

Soyabean-casein Digest
Medium
Ingredients Quantity for 1000 ml
Pancreatic Digest of casein 17 g
Peptic digest of soyabean
meal
3 g
Sodium chloride 5 g
Dibasic potassium phosphate 2.5 g
Dextrose 2.5 g
Distilled water Upto 1000 ml

Minimum sample size related to
Batch size
No. of items in the batch Min. No. of items
recommended to be tested
Injectable preparations
a) upto 100 containers 10% or 4 containers
whichever is greater
b) 101-500 containers 10 containers
c) >500 containers 2% or 20 containers
whichever is less
Ophthalmic and other non –
Injectable preparations
a) upto 20 containers 5% or 2 containers whichever
is greater
b) >200 containers 10 containers

No. of items in the batch Min. No. of items
recommended to be tested
Solids
a) <4 containers All containers
b) 4 - 50 containers 20% or 4 containers
c) > 50 containers 2% or 20 containers

For injectable preparations
Quality of each container Min. Quality to be used for
each culture medium
For liquids
a) <1 ml Total contents of a container
b) 1 ml – 4 ml Half the contents of a
container
c) 4 ml – 20 ml 2 ml
d) 20 – 100 ml 10% of contents of a
container
e) > 100 ml NLT half the containers
For solids
a) <50 mg Total contents of a container
b) 50 mg – 200 mg Half the contents of a
container

Test methods
 The test methods for the sterility of the
products are:
◦ 1. Membrane filtration method
◦ 2. Direct inoculation of the culture medium

Membrane Filtration method
 Appropriate for: (advantage)
◦ Filterable aqueous preparations
◦ Alcoholic preparations
◦ Oily preparations
◦ Preparations miscible with or soluble in
aqueous or oily (solvents with no
antimicrobial effect)
 Solutions to be examined must be
introduced and filtered under aseptic
conditions

Selection of the filters
 Pore size of 0.45μm
 Effectiveness established in the
retention of micro organisms
 Appropriate composition
 The size of filter discs is about 50 mm
in diameter

Procedure
 Sterilization of filtration system and
membrane filtration of examined
solution under aseptic conditions
(suitable volume, dissolution of solid
particles with suitable solvents,
dilution if necessary)
 The membrane is removed,
aseptically transferred to container of
appropriate culture medium

Scheme for sterility test by membrane
filtration

Direct inoculation of the culture
medium
 Suitable quantity of the preparation to
be examined is transferred directly
into the appropriate culture medium
 Volume of product is not more than
10% of the volume of the medium
 Suitable method for aqueous
solutions, oily liquids, ointments and
creams

Scheme for sterility test by direct
inoculation method

Interpretation of the results
 The culture media is examined during and
after incubation period to detect the possible
microbial growth.
a) The sample passes the test if microbial growth
is not found
b) If microbial growth is present, a retest is
performed. If growth is absent. Then sample
passes the test
c) If microbial growth is present in the retest also,
identify the organisms
d) If same organisms are found as in the first test,
then the sample fails the test
e) If different organisms are found, retest is
performed using twice the number of samples.
Passes if microbial growth is not found.

Pyrogen Test
 Pyrogens are the metabolic products
of gram negative bacteria
 Endotoxin – complex of pyrogenic
lipopolysaccharide, a protein and inert
lipid;
 Lipid part of the lipopolysaccharide is
the main pyrogenic agent;
polysaccharide part increases
solubility

Sources of pyrogen
contamination
 Solvent – possibly the most important
source
 The medicament
 The apparatus
 The method of storage between
preparation and sterilization

Animal and Equipment
 Selection of animals (healthy, adult,
NLT 1.5 kg)
 Housing of animals
 Equipment and material used in test
(glassware, syringes, needles)
 Retaining boxes (comfortable for
rabbits as possible)
 Thermometers (standardized position
in rectum, precision of 0.1oC)

Preliminary Test
 Intravenous injection of sterile
pyrogen-free saline solution
 To exclude any animal showing an
unusual response to the trauma
(shock) of injection
 Any animal showing a temperature
variation greater than 0.6oC is not
used in the main test

Main Test
 Group of 3 Rabbits
 Preparation and injection of the product
 Warming the product
 Dissolving or dilution
 Duration of injection: NMT 4 min
 The injected volume: NLT 0.5 ml per 1 kg and
NMT 10 ml per kg of body mass
• Determination of the initial and maximum
temperature
 All rabbits should have initial T: from 38.0 to
39.8oC
 The differences in initial T should not differ from
one another by more than 1oC

The result of pyrogen test:
No. of
Rabbits
Individual
Temp. rise
(oC)
Rise in group
(oC)
Inference
3 0.6 1.4 Pass
3 + 5 = 8 0.6 3.7 Pass

Limulus Amoebocyte Lysate
Test
 To detect or quantify endotoxins of
gram negative bacterial origin
 Reagent: Amoebocyte lysate from
horseshoe crab (limulus polyphemu or
Tachypleus tridentatus)
 Bacterial Endotoxin Test (BET)
 Monocyte Activation Test (MAT)

The endotoxin characteristics
 Thermostable
 Water soluble
 Unaffected by the common
bactericides
 Non volatile
 These are the reasons why pyrogens
are difficult to destroy once produced
in a product

Mechanism of LAL Test
 The test is based on the primitive
blood-clotting mechanism of the
horseshoe crab

Commercially derived LAL
reagent
 Bleeding adult crabs blood into an
anticlotting solution
 Washing and centrifuging to collect
the amoebocyte
 Lysing in 3% NaCl
 Lysate is washed and lyophilized for
storage

Procedure
 Test:
◦ Equal volume of LAL reagent and test
solution (usually 0.1 ml of each) are mixed
in a depyrogenated test-tube
◦ Incubation at 37oC, for1 hour
◦ Remove the tube – invert in one smooth
motion (180o)
◦ Observe the result

Different Techniques
 Three different techniques:
◦ The gel-clot technique – gel formation
◦ The turbidimetric technique – the
development of turbidity after cleavage of
an endogenous substrate
◦ The chromogenic technique – the
development of color after cleavage of a
synthetic peptide – chromogen complex

Gel Clot Technique
 A solid gel is formed in the presence
of endotoxins
 This technique requires positive and
negative controls
◦ Positive controls – a known concentration
of endotoxin added to the lysate solution
◦ Negative controls – water, free from
endotoxins, added to the lysate solution

Turbidimetric Technique
 The test is based on the measurement
of opacity change due to the formation
of insoluble coagulin
 Opacity is directly proportional to the
endotoxin concentration
 This technique is used for water
systems and simple pharmaceutical
products

Chromogenic Technique
 This is based on the measurement of
color change which is caused by the
release of the chromogenic chemical
p-nitroanilide
 The quantity of the p-nitroanilide
produced is directly proportional to the
endotoxin concentration

Advantages of LAL Test
 Fast
 Greater Sensitivity
 Less Variability
 Much Less Expensive
 Alternative to Animal Model
 More accurate than other
 Is performed in the pharmaceutical laboratory
 Specific for endotoxins of gram-negative origin
 Particularly useful for:
◦ Radiopharmaceuticals and cytotoxic agents
◦ Products with marked pharmacological or toxicological activity in
the rabbit ( e.g. Insulin)
◦ Blood products which sometime give misleading results in the
rabbit
◦ Water for injection where LAL test is potentially more stringent
and readily applied

pH measurement
 pH meter is used to
measure the pH of
parenteral
preparations.
 The pH of injectables
should be w.r.t pH of
blood (i.e., 7.35 to
7.45)
 The pH of
opthalamics should
be w.r.t pH of
lacrymal fluid (i.e.,
7.2 ± 0.2)

Parenterals
Problems Remedy
•Leakage of ampoules •Discard the ampoules
•Use new ampoules with proper
sealing
•Perforation in filter leading to
defective filtration
•Changing the filter

•Foreign particles/ dust Membrane filtration
•Fibres Optical and visual inspection
•pH Use of buffers
•Leaching Internal coating of glass container

References
 USP, Appendix 788, 56
 IP p 659 – 660
 Remington, The science and Practice
of Pharmacy, 21st edition p 1367 –
1374
 Michael J. Akers and Daniel S.
Larrimore, Parenteral Quality Control
 http://apps.who.int/phint/en/p/docf/

Ipqc tests for parentrals

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