Q There is a lot of emphasis now a days on 100% sampling

(all containers ). Is it necessary to do identification tests

for each drum / bag of raw material ? This could pose a
problem in storage of materials procured for long term
use, expecially if the material is deleterious.

A Currently, the recommendation, is that every container of an

active material and certain critical excipients should be
tested container wise. The logic behind the requirement is,
that many users have found, variations in quality parameters
from container to container due to either inadequate
uniformity in the lot material or, due to exposures to
temperature and humidity changes during transportation.
Sometimes, a shipment of raw material received has a
number of containers, which represent an inventory for use
over a period of 3 to 6 months, especially in the case of
improved materials.

Some materials are very susceptible to moisture and

therefore, once the original container is opened, it must be
used as soon as possible. In such cases, it is suggested
that, a limited number of containers should be sampled as
a sublot. Based on the test results, the lot would be either
accepted or rejected.

Further, the remaining for would, from time to time, be

divided into sublots and tested, depending upon the usage
plans. In the event of a lot being accepted, and the goods
paid for, but, if subsequently a container does not meet
specifications, it should be returnable to the suppliers.

Q Is it advisable with reference to GLP, to have different

sampling schedules for active and inactive raw materials ?

A Both, active materials and excipients have to be sampled

according to the same sampling plan, 1 + n containers.
Active materials would need additional sampling to ensure
quality compliance.

Written SOPs for sampling plans and procedures are

required for cGMP compliance. Dr. S.N. Lyer’s previous
answer explains briefly, how sampling of active and
inactive excipients may be done.
Q Why should sampling be done on bulk as well as finished
products ?
A The bulk product samples drawn from different dosage
forms could have some limitations on the sample
uniformity. For example, the bulk product is tested and
released for packaging. But, because of change in
requirements or other considerations, remains unpacked for
long periods of time, or, in situations when part of a bulk
lot is packaged at one time and the remaining, after some
time, deviations could occur. It would be appreciated that
bulk product storage conditions do not replicate that of a
finished batch. Therefore it is desirable that the finished
product batch also be tested to its release for sale.

Q If the expiry date of raw material is 5 years and it is

used in the last month before expiry for manufacturing the
product, what should be the expiry of finished product ?

A This question has been raised from time to time in various

forums. The expiry date to be given on the product
depends on the stability data that the manufacturer has of
the product. In the event of the raw material being used
in the formulation nearing its expiry date, the assignment
of expiry date would depend on the stability data the
company has, of experimental batches made under identical
conditions. The drug and cosmetic rules provide, that the
licensing authority may accept any expiry date, so long as
it is supported by stability data. It may also be noted,
that if a raw material carries a 5 year expiry date, its
formulated dosage form need not necessarily have a 5 year
life.

Q Is there any sampling device, other than rods and scoops

which can collect samples from different depths of the
container ?

A A thief sampler, which is a hollow cylinder cut into half

and provided with a pointed end could be used. Still
better, is two hollow cylinders, one fitting into the other.
An opening is provided in each of these at the same depth.
A tapered end is provided. The cylinders are inserted into
the bag / drum upto the desired depth. Making sure the
openings of the inner cylinder are diagonally opposite to
that the outer cylinder. After the device is inserted to the
desired depth, the inner cylinder is rotated by 180, so that
the two openings are adjacent each other.
A slight left to right movement of the device will draw
the powder into the inner cylinder, which is then rotated
 again by 180 and the device is pulled out. This way, the
samples from different depths can be collected.

Please elaborate on the retest period for active excipients ?

A Normally, the retest date should not be more than a year

for anything. For items having volatile components, for
example, flavours, a retest period of 6 months is
recommended.

In the faculty’s opinion it will be very poor inventory

management, if there is need to retest any material more
than once, although, this is permissible within the full shelf
life of the substance.

Proper planning of production will aid procurement

procedures and avoid retesting.

Q For every market complaint received, is it necessary for the

manufacturer to do the analysis of product from a very
limited retained sample ?

A No, it is not essential that the retained sample be fully

analyzed or even partly analysed for every market
complaint received. For example, if the complaint refers to
shortage, empty pockets, broken bottles, the potency
analysis of the product is not going to tell us anything
about the complaint. Hence, what investigation, if any, is
required depends on the nature of the complaint.

Q In the current scenario of IT/ Kanban, reduction in quality

inspection and quality at source, is currently practised,
especially by Engineering Industries. How much of this is
applicable to Pharma Industry ? How can we implement
these at minimum cost ?

A JIT or Just In Time inventory management is a very

laudable objective to have. However, the current scenario,
especially for Pharmaceutical Industries, where several raw
materials are imported and others are obtained not
necessarily from one vendor on a need basis, this is not
practical. If the company has a selected, reliable vendor
who can give assured quality at the specified time, JIT can
be instituted.
Q What are the pre entry cleaning guidelines for various
types of packings HDPE bags, fibre drums, jute bags,
Cardboard boxes ?

A Pre entry cleaning procedures for various materials received

in the warehouse, should have cleaning techniques
appropriate to the type / constitution of the outer container,
the surface of which is to be cleaned. The level of
cleanlines required, has to be determined by the company
depending on the nature of raw material, outer container
pack and the handling conditions in the dispensing area. A
system for validating such cleaning would be desirable.

Q What is the retests period of raw materials which has

microbiological testing in the specification ? How many
times should this material be tested ?

A The retest period for a raw material is usually assigned on

the basis of either experience gained within a company six
months or one year being the norm or sometimes the
expiry date of material is specified by the manufacturer.
The objective of retest is to ensure that during storage, the
material has not undergone any change in its properties.
Those materials requiring microbiological testing, should
be tested for microbial content along with other parameters.
However, in the event of raw materials showing a
microbial contamination of increased CFU, it would be
desirable to test stored material at relatively shorter
frequencies, than that meant for chemical retests. This is
just to make sure that the microbial population is not
increasing.

Q Why do granules sometimes get hardened during the

granulation process ?

A Many factors influence granulation, some of the common

cases for formation of hard granules are :
* The wrong choice of a binder, such as binders with high
adhesive property.

* High binder concentration.

* Use of excessive granulating fluid or over wetting of the

powder.
* When the end-point of granulation is not properly
ascertained.

* Use of water as a binder.

The bulk density of material used in formulation varies

from supplier to supplier and hence, the quantity of water
added also changes, leading to the formation of hard
granules.

Q Batch numbering and code numbering systems vary from

one company to another, as they are designed in house.
Can you recommend, from experience, a system that you
consider the best ?

A There is really no system which can be considered the

best. A system must serve you well at the time you use
it, and it must be constantly reviewed to see if the
purpose its being achieved. In my experience, the easiest
method of batch numbering is the straight forward serial
number system for each product. Complex systems including
alpha prefixes and extra numberals to know the product
identity, year and month of manufacture etc. Serve no
purpose, because, all this information has to be provided
on the labels in any case, as per regulatory requirements
and there is no point in complicating the batch numbering
system.

Code numbering of printed packing, materials is a different

proposition and there are advantages in coding. Here, I
would recommend an alphanumberic system, built around a
unique code number for each product, with suitable prefixes
and suffixes to identify the packing material and label
copy. For example, a label for a product with an assigned
code number say 345 could be coded as L 345 Al where L
would indicate that it is a label and A1 would refer to the
specific and current printed copy matter on the label. If
this matter were to be amended the code would be changed
to L 345 A2. Likewise, if the pack for the same product
were to contain a printed carton, it could be coded as
C345-B1 and with any changes. In copy it could change to
C345-B2 and so on.

Q Sampling plan for packaging materials - should it be as per

ISI standards, or can it be changed on the basis of the
vendor ?
A Sampling plans as laid down in the ISI standards may be
followed, if the vendors are prepared to work out the
AQLs with the buyers. In the absence, an agreement it
will have to be on the basis of what the vendor is
prepared to accept.

Q Microbial limit test for FP is not mandatory as per IP

Schedule M mentions that all pharmaceutical preparations
should be free from pathogens. Are all batches of FP to
be tested, for the absence of pathogens ?

Should this test be used as a release criterion or, is

monitoring enough ? Also, should a total microbiol cannot
be done on FP ?

A It is essential that all non-sterile preparations be subjected

to controls for microbial contamination. For this purpose,
the source and nature of contamination of all the products
should be ascertained by routine monitoring of both, the
input mateials and the products, as well as, the
environment. In which the products are manufactured. It is
only after this has been done, that microbial limits can be
formulated and then enforced, either as release limits or, as
monitoring criteria. The WHO has recommended limits for
various types of materials and products Reference may be
made to them for guidance.

All questions so far have been answered by R.S. layer,

Senior Faulty. The Academy.

Q Please suggest what type of LAF workstation should be

used where basic filling / stoppering and sealing operations
are carried out for sterile products.
A Vertical Downflow Airflow Unit is generally the
recommended LAF work station for the operation.

Q Which is the most suitable validation technique for

autoclaves and dry heat sterilizers , Chemical Indicators or
Biological Indicators.

A Firstly, temperature is not the parameter that needs

verification.

Chemical indicators are normally melting pellets, color

change strips or witness tubes. They measure steam
 penetration only and not the quality of steam i.e. Dry
Saturated Steam. They exonerate the vacuum cycle.

Biological Indicators make use of thermo resistant bacilli.

These indicators, besides measuring steam generation, also
show the enthalpy transfer to the bacilli and give an
indication of the temperature hold. TME as an integrated
event. Strips of B. Stearothermophilus spores are used as
the challenge indicator when moist heat is used.

In dry heat sterilization, spores of B. Subtilis sub species

niger is the preferred indicator.

Here is a quote from the Encyclopedia of Pharmaceutical

Technology Page 27, Vol 2. ( Swarbrick and James C
Boylan ).

Two primary uses for B1 are validation and monitoring of

sterilization procedures. In validation. Bls are used to
ensure that a particular carefully controlled process will
produce a sterile product under conditions of maximum
challenge ( i.e. worst - case conditions ). In monitoring, Bls
are used to ensure that this process remains capable of
producing sterile products, even under worst case conditions.
The advantage of Bls and an attribute that cannot be
duplicated by any physical measurement is the ability to
integrate the lethal effect of sterilization agents under
conditions in which at least two parameters are being
varied at the same time. This makes the determination of
the effectiveness of the sterilization processes, so that
validation can be carried out as well as continuous
monitoring. The more information can be found in the
Academy’s Update on the subject.
Q What is the ideal gap required between filled volume and
the space above the liquid upto the constriction of the
body of the ampoule ?

A Ampoules would most probably be terminally sterilized by

moist heat after filling. During this process the temperature
rises from 20 C to 121 C . This would mean an increase of
the liquid volume by 6% and so a minimum head allowance
of 6% should be maintained.

In the case of vials, compresion of head space air ( or

nitrogen ) should not displace the neck of the stopper
within the throat of the vial to an extent, that the stopper
 pops out. The seal force must not be overcome by the
compressed force.

Q What would be the method of validation of an autoclave

size 4’ x 4’ x 6’ with load of 515 ml. At 15 psi, 121 C for
20 minutes to avoid pocketing of steam in the chamber,
Please suggest the ideal validation method and how
frequently it should be done?

A An autoclave that uses direct entry steam as the sterilizing

agent, will always have the problem of pocketing of steam
in the chamber. The answer lies in whether the autoclave
has a purge system or not ? The purge system does not
guarantee the absence of pockets, but definitely voids
pockets sufficiently to dramatically reduce the Cold slots.
Validation can be done by either the 12 probe method or
by the Bacteriological method, but it can be established
only after the process cycles are understood. Today’s
technology makes use of super heated water as the
sterilizing agent to avoid the problem of steam pocketing.

Q Why should unlubricated granules not be kept in drums

similar to lubricated granules ?

A Unlubricated granules should not be stored in similar

containers to avoid the mixups between unlubricated granules
and lubricated granules. Even the labels used should be of
a different color to facilitate easy identification by even
the illiterate workers.
Q If a drug is non- pharmacopial, how do we select the
dissolution conditions media composition agitation ( rpm )
and USP apparatus I or II for dispersible tablets ?

A If the drug is non pharmacopial, the medium selected

should be water N/10 HCl or a suitable buffer upto pH 7
USP Type 11 apparatus should be used and agitation
should be 100 rpm.

Q What should be the correct head space in a LVP bottle ?

How much more than 6% by volume ?

A About twice, 10 to 20% The insights dont’s end there.

Full up. Bottle volumes is usually 20% over labelled dose.
 At 121 C, a bottle filled at say, 20 C wet bulb room
temperature, has two internal pressures.

1 Partial vapoure pressure which changes from 0.025 to 0.050

bar, and.

2 Partial air pressure which rises from 0.988 to 1.330 bars

(applying perfect gas laws to air ).

The total pressure of 1.38 bars means, that the stopper is

pushed at about 1.4 kgs per sq. Cm.

Standard bottles used in our country, 250 ml and up, have

a neck of 22.5 mm. That is a force of 5.5 kgs exerted
the explusion thrust. Residual stopper sealing force is
always of concern and to be kept in mind when adjusting
head space.

Q Why is steam, with all its associated problems used for

sterilization ? Why not super heated water, as is used for
stopper ?

A The answer to this, has to do with specific enthalpy. At

121 C, water is at 500 Kilos joules per kg. Associated
with steam, its at 2530 Kj per kg. And, evaporation
enthalpy is at 2022 Kj. This accounts for the large heat
transfer capacity of steam and its ability to penetrate the
load mass getting to where the bacteria are resident.

With stopper, the position is that steam should not be

allowed to penetrate as it destabilises the interstitial,
intercrevical matrix of the closures. Also these elastomers
vulcanizites are prone to regression by the action of steam.
So it is prudent to use high temperature water for closure
sterilization.

Q What is superheated steam and what are its limitations

when used in an autoclave ?

A It is important that a critical balance of pressure and

temperature be maintained in an autoclave, to ensure a
contineous supply of dry saturated steam. If the temperature
rises or the pressure drops, the degree of saturation is
reduced and the steam becomes superheated.
 The objection to super heated steam is technical, enthalpy
transfer. Superheated steam inhibits the transfer of moisture
as it condenses less readily than dry saturated steam. Due
to decreased condensate levels, the moisture required to
initiate cell destruction is just not available. The cite just
one value here the specific volume of steam at 126 C is
0.743 m3 / Kg and at 121 C is 0.841 m3 / Kg nearly 13%
less at the higher temperature.

Q What happens to the F value when you do different hold

times ?

A The F sub zero is also called F physical, because it is a

parametrized process equivalent time at the reference
temperature of 121 C with Z taken to be 10 C.

If you do a Log 6 overkill, that value is OK. But in

reaily we work on retained bioburden with different
decimal reduction times. Then, we have to arrive at a F
and Z by T value.

Q What happens when the steam quality supplied is wrong ?

What happens to validation ?

A That’s an operating conditions change and mandates

requalification of process.

Validation has to be maintained within the limits that are

documented. Change control, which in itself, is a written
procedure, and outlines the analysis and the implementation
of change. This often necessiates repeating all elements on
the validation studies.

Its not only steam quality that demands change control. Its
venting, product loading, and, density, even changes in
process control software, attract the full force of change
control.

Q How often is requalification necessary if nothing goes

wrong .

A Usually, once in a year, assuming consistency in operational

procedures and SOPs in calibration, maintenance, inprocess
parameters, in process audits in other words, every activity
that relates to end product assurance.
Q Why it is necessary to establish retained bioburden ?

A The US FDA says to describe and submit a procedure for

routine monitoring of bioburden to ensure that established
limits are not exceeded.

The industry position is that with log 6 overkill, there

really is no need for this. Also where there is no substrate
for microbial growth like a container, or inert items like
closure, such a program is not called for.

Q What are the types of air samplers in use and what are
their disadvantages ?

There are various air samplers that are available. The

popularly used are :

(a) Cascade sampler :

A stack of agar plates is separated by perforated plates the

perforations decreases in dimension, going from the top to
the bottom of the stack. A sampling rate of 1.0 ft3
minimum, is maintained. Microorganisms attached to the
larger particles, impact on the top agar plate and those
attached to smaller particles, cascade over, through the
perforations, until they impact on a plate lower down in
the stack. After the specified sampling time, the plates are
incubated and examined.

(B) Slite to agar sample :

The air sample is drawn through slits and impinges on a

rotating agar plate. Minimum impingement of the
contaminants in the air sample is achieved by controlling
the dimensions of the slits, the distance from the slits to
the agar surface and the air flow.

(C) Single sieve to agar sampler :

The air sample is drawn through a perforated disc and

impinges on a 55 mm agar contact plate.

(D) Centrifugal sampler :

The air sample is drawn onto the sampling head by means

of an impeller. The impeller then directs the air onto an
agar strip filled around the circumference of the sampling
head. The air sampling is done for a specified time period,
after which the agar strip is removed and incubated.

(E) Filtration :

The air sample passes through a membrane filter for a

specified time period. The filter is then incubated on an
agar plate.

(F) Liquid Impingement :

The air sample is drawn through an aspirator bottle

containing a buffer or any other suitable medium. After the
specified sampling time the solution is examined for the
presence of microorganisms by a suitable method.

Each of these air sampling devices have several

disadvantages.

The cascade sampler has a very low sampling rate to study

the most critical environments.
Only some of the slite to agar samplers can take a
statistically large enough samples ( 3.0 - 3.5 m3) to enable
the examination of critical environments. They are also
rather cumbersome to use. Procedures for operating these
samplers at higher sampling rates / longer sampling periods
need validation.

The efficiency and sampling rate of the single sieve to agar

sampler is even lower than that of slit to agar samplers.
The centrifugal sampling device is the most convenient use
but there are so many uncertainities surrounding its
sampling rate, that data interpretation is more difficult.

The filtration technique can take statistically large enough

samples but again the process needs validation.

The liquid impingement method introduces additional

preparation procedures for the aspiration bottle and buffer
solutions and procedures for recovery of contaminants. Care
should be taken to prevent spillage of buffer solutions in
the critical area.
Q How many media fill tests would be required to establish /
validate an aseptic processing operation ? How often should
these tests be performed ?

A There are no uniformity accepted criteria which indicate

how many tests establish this procedure. But, Yes, what is
accepted is that more than one media , fill test is required
to evaluate a new processing operation. There are
suggestions that three media fill tests should be run for
the initial validation. But, if three tests provide divergent
results it gives less assurance than two consecutive
acceptable tests. What is important is that the number of
tests should be based upon sound scientific and technical
judgement rather than relying on any specific number of
tests to be done.

Revalidation :

If no changes have been made to the manufacturing

process and the media fill test data and environment data
consistently meet specifications, then there is no need to
perform more than one media fill test in a year, for each
aseptic processing operation :

However, the revalidation schedule should be based upon an

evaluation of process changes, historical media fill test data
and environmental monitoring data.

Q In general,, potent medicaments are not added during the

dry mixing stage. These are generally added at the
lubrication stage. Will this product pass the content
uniformity test ?

A If possible, it is advisable to add a potent drug in the dry

mixing stage itself, to ensure uniformity of mixing.

Whether the drug is added in the dry mixing or lubrication

stage, the process should be validated.

The validation of the process would ensure uniformity of

content, even if the drug is added in the lubrication stage.

Q It is noted that permeability of gases into formulations is

much higher in plastics. Will this affect the shelf life and
efficiency of drugs as compared to similar formulations in
glass bottles ?
A From the point of permeability, glass is definitely less
permeable to gases when compared to plastics. Hence, the
shelf life of formulations stored in these containers will
vary.

Photosensitivity is another point to be considered. The

plastic let in a totally different wavelength of light.

But, it should be remembered that glass has the

disadvantage of surface alkalinity and leaching of
extractives.

Permeability is only one of the factors which affects the

shelflife. The self-life of a product in any pack depends on
many factors, all of which would have to be examined
before deciding between glass and plastic. These factors
should be ascertained at the time of stability studies and
an indepth study carried out. The effect of the container
should be factored in, before fixing the shelflife of a
product.

Q What is the maximum limit to store tablets in one single

container ?

A By the words maximum limit I consider time limit and

would say that, every organization has different standards,
depending on convenience.

Let us take an extreme case.

A batch of tablets is packed in a bulk pack of thousands
and sent to the market with a shelf life of say, 3 years
that the tablets can be stored in the container for 3 years.
Many companies are exporting bulk pack of 5,000 or
10,000 tablets, which means that the tablets are stored in
the container for 3 years.

Extending the same logic, can it be considered safe, if

bigger bulk packs of tablets can be left on the shop floor
for 3 years ? Before we answer this, please consider
hermatically sealed containers sent to the market vis a vis
tablets stored on the shop floor.

If we have to set time limits for storing tablets in a

container, then one has to set limits for storing granules
 before compression, and also a limit for storing recovered
tablets before processing.

My personal view is that, time limits for keeping tablets

should not exceed more than 3 months. Otherwise, it
indicates bad planning, on part of all the departments of an
organisation. Instead of keeping these tablets as such, it is
always better to keep these as a final pack because, then,
it is in a hermatically sealed pack.

Q What could be the best system for pest control in

production manufacturing areas, where partitions are made
up of wood / plywood ?

A Please pull down the entire structure and go for aluminium

and glass. No pest control should be done in production
areas. There is no guarantee that pesticides are not carried
into the products.

Q Is it necessary to boil the sugar syrup in liquid products

to 70 0 C - 80 0 C ? Why ?

A Most of the manufacturers make sugar syrup by the hot

process. But, there are a few who make it by the cold
process. Sugar is one of the ingredients which is bought in
gunny bags by most of the manufacturers and so, it is
always safe to make the syrup by the hot process. If one
has to make this without heating, the parameters to be
considered are : what is the concentration of sugar going
into liquids, what is the quality of sugar and foreign
matter, how long will the liquid be kept under storage till
the final product is packed, how is it stored and whether
the product can withstand the preservative challenge test.
We also should consider what is the quality of sugar over
a period of one year and the quality of the syrup made
with the same.

It is essential to validate your process and product fully

with respect to the above mentioned points and new
materials going into the liquids. If you are confident and
can assure the quality, you may use the cold process.

When the hot process is used, sugar syrup is prepared at a

temperature which is around 65 0 C and has a concentration
of sugar, which is well above 60%. This will certainly
prevent the growth and also kill many microorganisms.
Q Please describe the storage conditions, dispensing procedure
and sampling procedure for cytotoxic drugs.

A For most of the raw materials, storage conditions and

processing ( manufacturing ) conditions are the same. A few
materials, whose initial activity is likely to be affected by
higher temperature, are kept at lower temperatures of 2 to
5 0 C. Please take this into account before deciding storage
conditions of cytotoxic drugs. There has to be an exclusive
dispensing area with reverse laminar flow, and depending
on the hazardous and potent nature of cytotoxic drugs,
proper clothing, hood and mask should be provided, while
sampling. This can also be sampled in the production area,
like sterile raw material sampling.

These questions have been answered by I.R. Thamankar

G.M. Formulations Kopran

Q What is Dutch Mesh of fluidized bed dryer ? Does it differ

from conventional mesh for sifter ?

A The Dutch Mesh used in fluidized bed dryers is also known

as Hollander Weave mesh. Twill Mesh or Multi mesh. This
twill mesh is stronger when compared to the ordinary
mesh. Owing to its durability, it is more commonly used
in fluidized bed systems as compared to the conventional
mesh used in sifters.

In the Dutch weave mesh, there are two different guages

of wires used in the warp and weft of the wire screen,
thereby imparting much more strength and durability to the
screen. It therefore lasts longer and also withstands higher
pressure as compared to a conventional mesh. The twill
mesh also cleans more easily.

Q What’s is the material of construction of FBD bags ?

A Normaly, FBD bags consist of a fabric containing a mixture

of polyester and cotton in equal proportions. Premium filter
bags are generally made from epitropic cloth, which is
essentially a carbon impregnated fabric. While the material
cannot be considered to the antistatic, it is at least
conductive, so as to drain away electrostatic charge.
Q How do you protect punches from rusting when it is not
permissible to use kerosene ?

A After compression, remove the powder adhering to the

punches using a dry cloth. Then, soak the punches for five
minutes in warm water which contains 20% detergent and
clean them with a nylon brush. The punches are then
dipped in plain water and wiped thoroughly using a dry
cloth.

A small amount of lubrication oil is then applied, at the

tip and other parts of the punches before storing them in
the punch trolley or the punch box.

Q How can we reuse enteric coated tablet rejects ?

A A maximum of 2% of the rejects may be reused by adding

them to a fresh batch.

The enteric coating part of the rejects is washed using

acetone. The core tablets are them dried. After drying, the
tablets are crushed. The crush is sieved through a 60 mesh
sieve and the content / gm checked. The contents are
calculated as per assay and added to a fresh batch at the
time of formulation.

Note :

It is not advisable to reuse the enteric coated rejects of

highly hygroscopic tablets like sodium valproate

These questions have been answered by K.P. Lucasmani,

Manager Production, Sun Pharmaceuticals, Chennai

Q What should be the acceptable tolerance limits for vitamin

B12 and folic acid content in multi-vitamin tablets or
capsules ?

Are formulations of multi-vitamins specified in any of the

Pharmacopoeias ? What should be the minimum limit of
these drugs during their Shelf life period ?

A Some formulations, mostly tablets and capsules are official

in the Pharmacopoeias. In the USP, preparation of both,
water souble and fat soluble vitamins are official. These
include Decavit, Hexavit and Oleovit which contain vitamin
 A.,D, calcium, pantothenate and the Bcomplex vitamins.
There are no official products prescribed in the IP. If the
product is official, then the limits should be as per the
pharmacopoeia. Otherwise, they should conform to the
Schedule V of the Drug and Cosmetic Act. 1940 This
schedule lays down the standards for patent or proprietary
medicines.

The standards prescribed do not apply to preparations

containing a single vitamin only and to any preparation
containing vitamins for parenteral use. These limits are
relaxed for patent and proprietary preparations containing
vitamins intended for the treatment of specific conditions
or diseases. In such cases the Licensing Authority may
permit the addition of vitamins, if satisfactory evidence is
produced to justify such a relaxation.

For preparation containing vitamins, only a lower limit of

90% is applicable. That is, the content of active ingredients
in a vitamin preparation shall not be less than 90% which
is also the shelf life of the drug.

The dosage limits of vitamins B12 and folic acid are given
in the Table below.

Q In the case of a loan license manufacturers, for how long

should the records of manufacture be retained by the
manufacturer ?

A The records retention requirements for a loan license

manufacturer, that is a contract manufacturer are the same
as that for a principal manufacturer.

The retention time for all classes of drugs is for five years
after the date of manufacture.

The loan licensee should posses all documents records

pertaining to the manufacture of a batch, so that a complete
product history of the drug is maintained. The loan licensee
may enter into an agreement with the manufacture to the
effect that all the records will be available with the
manufacturer. The records must then be kept at the
manufacturer’s facility. If they are maintained elsewhere, it
should be possible to retrieve them immediately by
computer or other means.
Q During repackaging, is it allowed to allot a single batch
number, to two or more mixed lots of bulk finished product
repacked during the same run.

A Allotting a single batch number to a mixed batch of the

finished product, even if the records clearly indicate and
identify the batches that are being mixed, would cause the
finished repackaged product to be a violation of cGMP
norms.

Identifying problem lots or batches in the event of a

complaint or finding of a defective product, assigning
appropriate expiry dates, and collection of representative
samples may be a problem when batches are mixed
together.

These questions have been answered by Mr. M Kannan,

Senior Drug Inspector, Drug Control Dept. Chennai.

Q How important is testing for microbial contamination in

non- sterile products ?

A cGMP regulations require that there should be written

procedures for prevention of microbial contamination in
nonsterile products. These should ensure that there is no
contamination or growth of organisms in a product. The
prevention of microbial contamination should be evaluated
on a case to case basis, depending on speciation, number
of organism type and use of dosage form, route of
administration etc.

Microbial contamination in a product could adversely affect

its stability, react with or, damage the container system,
interfere with analysis affect bioavailability and pose a
threat to efficacy of treatment. It is also important to
establish production time limit to prevent microbial
contamination. Time limits for one completion of every
phase of production must be established and followed.

Q. According to cGMP is it necessary to have different

departments for the Cephalosporins and Penicillin
formulations ( dry powder Injections ) ? If so, why when
they are of the same B-lactam group ? Also, please let us
know whether these departments should have separate
building or block as per the U.S. FDA ?
A. An anaphylactic reaction to penicillin or for that matter to
cephalosporin - can be life threatening.
In the U.S. regulations for control of penicillin cross
contamination came into effect in 1969. About that time,
methods for detection and quantifications of residual B-
lactam antibiotics were evolved by what is now known as
NCAA ( National Centre for Antibiotic Analysis ).

Those methods, now outdated, gave a LOD (Limit of

Detectability ) of 5 ppm for cephalosporin and, near zich
for ampicillin. Naturally, that inadequacy combined with the
gross detection by the FDA and the Industry of penicillin
in non - antibiotic preparations let cGMP to mean
segregated manufacturing facilities.

The microbial analytical approach gave so specifically in

the analysis of drug residues, until Herbst in 1975
proposed the Bioautographic method that give a LOD of
0.5 ppm.

Dr. Meera, besides detection, a vital requirement for cross

contamination control is your ability to inactivate the
contaminant.

Please refer to Methods in Enzymology Vol. XLIII

Antibiotics Academic Press
Many different enzymes are capable of catalyzing the
hydrolysis of the B Lactam ring, and that, B-Lactamases
from at least 25 different strains of bacteria have been
purified and studied. Since B-lactamase preparations are not
standardized and are available from many different sources,
care must be exercise by the laboratory to employ a
suitable B-Lactamase preparation of sufficient titer, to
provide proper inactivation under specified analytical
conditions.

The B-Lactamase inactivation of the test solution may

produce the following results :

1) Complete inactivation (a positive test ) : The loss of

activity of a treated versus an untreated aliquot of the
sample solution indicates that a B-lactam residue was
present. A quantitative determination of the residue can be
made in terms of an appropriate standard.
2) Incomplete inactivation ( a presumptive test ) : A significant
reduction in the activity of treated versus untreated aliquots
of the sample solution indicates that a B-lactam residue
was present, but that other active agents or ingredients
have caused interference problems. Accurate quantitation of
the B-lactam residue is not possible under these conditions.

3) No activation (a false test ) : No significant reduction in

the activity of treated versus untreated aliquotes of the
sample solution indicates that interfering agents or
ingredients may be masking the presence of a B-lactam
residue. No quanlitative or quantitative analysis can be
made of the residue.

4) No activity ( a negative test ) The absence of zones of

inhibition from treated and untreated aliquots of the sample
solution indicates that no B-lactam residues has been
detected ( within the limitations of the test method )

Now comes my opinion not an answer. Separate AHU

makes sense. Separate block, better sense, separate building,
best sense ?

Normally, is our LAF’s the Air Velocity should be 90 +

20 CFM. During validation. I found out that some of the
HEPA Filters located in supply duct of class 1000 Area,
were having Air Velocity of 250 CFM, which seems to be
quite high and detrimental to HEPA Filter as well as
aspectic area. Kindly comment. Also, what are the possible
ill effects of such a type of system ?

Rajiv, if you don’t mind, I will somewhat broad base the

reply.

Laminarily, or lack of turbulence, is the design intent of

all systems where one works in ciose proximity of a HEPA
futer.

For horizontal use, air velocities upto 0.7, m/sec the upper
threshold are fine, provided you have linear containment of
the airflow, like a canopy, sides, and a table. Values must
respect cross section aspect ratios.

For vertical airflow :

 With gravity as a natural vector this value goes down to
0.4 m/sec. And, the U.S. Industry tends to 0.35 m/sec.

The new standards have dropped the description LAMINAR

for the more apt Undirectional, and, give no limits for
velocity, implying that laminarity is case determined,
calling for a one on one analysis of User needs. It cannot
be brochure specific.

Coming to the other applications. Terminally mounted

HEPA Filters are called mixed flow systems the volumes
of air is predicated by the number of air changes per hour
in the clean room. That is the criteria. Consequently, the
airflow is designed to be maximal for that particular filter
size. The threshold velocities are 1.5 m / sec. Which is the
usual rating for a 2000 m3 / Hr capacity HEPA filter.
Capital and operational costs are minimal.’

In aseptic processing areas that velocity is too high, as

intra zone over pressure control cannot be ramped. It is best
to stay with 0.75 m/sec Operational costs are less, but,
capital costs are inversely proportional, while functionally,
the area performance is directly proportional.

The other ill effects of higher exit face velocities are the
bellow back within the HEPA matrix, particularly, in
exchange areas. That assists grow through conditions as
until recently terminally mounted HEPA were made to filter
air at near dew point. Micro droplets excasebrated lead to
higher plate counts during the monsoons.

Those problems have, of course, been fully overcome today

with the new enhanced performance HEPA Filters.
These queries have been replied by B. Singhania.

Do we have special coated punches and dies for special

toolings are available for compressing materials containing
salts or products that cause rusting. Some options are a.
High Chrome High Carbon ( HCHC ) steel can be used to
minimise rusting. As this steel is brittle, punch cavity
should preferably be selected as shallow concave to prevent
chipping of edges ;

h Oil Hardened Non Shrinkage ( OHNS ) Steel for punches

with hard chrome plating can prevent rusting upto some
extent :
c Hardenable stainless steel can be used for punches, but
compression pressure has to be restricted. However, this
steel is not available indigenously.

Types of Breaklines ( A-G) are they followed by all tool

manufacturers as standard ?
Dust cap : cleaning after compression - how convenient is
it ? Regrinding of punch tip : is it possible for the
monogranted punches also ?

Types of Breaklines ( A-G) are not yet followed by all the

tool manufacturers in India. However, they are followed as
standards in European Countries.

Dust Caps :

Two types of dust caps are used to keep upper punch dust
free, so that cleaning frequency of upper punch is
minimised.

Type A :

Elastomeric Caps fined on the upper punch tip. In this

type the upper punch shank is prevented from dust
marginaly (25%). Individual Rubber cap has to be procured
for different size of table.

Type B :

Elastomeric seat and “rubber” bellows are fitted on upper

punch shank. This system prevents dust from reaching the
punch shank to a great extent (80%) Upper punch shank
has to be modified so that “rubber” seat can be fitted.

Common rubber seat and bellow are used for all size of
tablet, available indigenously for “B” tooling.
Regrinding of punch cavity is possible only in round,
concave cavity without monogram. An existing monogram
can be removed by regrinding but not vice versa.

Q Wha is impurity profile ?

Sipali Pharmaceuticals

A An impurity profile is a list of all possible related

substances that could be present in a compound.
 The USP defines an impurity profile as a description of
the impurities present in a typical lot of drug substance
produced by a given manufacturing process.
This standard profile is developed early on during the
developments and stability studies of a pharmaceutical
compound and a record should be maintained of this
exhaustive study. Each commercial lot of a compound
should be comparable to this record. This profile could
also be called the reference profile. The Quality control
department would refer to this profile for assessing the
purity of a batch of an active ingredient and when
evaluating any process changes.

An impurity profile should give certain basic information

of impurities present at or above the 0.1% level or
sometimes even lower depending on the toxicity of a
compound.

The information includes :

* Test of identity
* Ranges normally found
* Limits
* Description of the impurity

Whether it is an in- process decomposition product,

unreacted intermediate, etc.

The methods developed for detecting impurities should be

appropriately sensitive and capable of detecting and
quantifying the impurities.

At least two methods ( e.g. HPLC, GC ) could be used for

the routine purity testing of a pharmaceutical compound.
Identifying and developing an impurity profile for an active
pharmaceutial susbstance is consistent with cGMP.

Q Should active ingredients be tested only for those

impurities stated in the Pharmacopoeial monograph, or must
batches be sufficiently characterised for purity ?

A The monograph in any pharmacopoeia cannot list out tests

for every impurity, contaminant ( including microbial ) or
adulterant that might be present. These could arise from a
change in the source of material, changes in processing or
 from an extraneous source. Tests suitable for detecting such
substances should be employed in addition in the tests
provided in the individual monograph.

The USP supplement V ( official Nov. 15, 1996 ) states

that for active Ingredients major impurities ( 0.1% or
greater) which are not listed in the product monograph and
cannot be reliably analysed by its methods should be
named, quantified and included in the certificate of analysis
of the official substance. For those impurities for which
toxicity is a concern a lower qualification threshold and
more stringent specifications should be prescribed.

Q What should be done with a raw material which conforms

to all the standards laid down in the respective
monographs of IP / USP / BP, but gives positive tests for
other types of impurities which are not mentioned in the
monograph ?

A If the impurity is not mentioned in the official monographs

and in the company’s internal specifications, it is legal to
go ahead and use the raw material.
None the less, having detected the impurity, it is essential
that it should be quantified and investigated with the help
of literature. If it is ascertained that there might be any
kind of adverse effects because of the impurity, it must be
put on hold.

But, irrespective of whether it causes, adverse effects or

not, the presence of such impurities should be taken up
with the supplier of the raw material and steps taken to
ensure that these additional impurities are eliminated.

Q In formulating a batch to provide not less than 100% of

the labelled or established amount of active ingredient, most
firms consider such ingredient attributes as assayed potency
and water content ?

A It is an essential of GMP to consider assay, values, water

content and any other factors that might affect the potency
of batch during formulations.

If a batch is labelled 100% but the active ingredient is

known to contain about 5% of moisture, the effective
potency of the formulation is 95% And it is also possible
that there might be variation, depending on the actual
 moisture present. So, we have batches which contain 96%,
97%, 95% and so on, of the active ingredient. Establishing
a shelf life becomes complicated

So, the moisture content should be factored into the

calculations to ensure that all batches contain 100% of the
active ingredient.

Q When the labelled expiration date states only the month

and the year, does that mean that the drug expires at the
end of the specified month ?

A If the expiry date states, for, e.g. Feb. 1998, it means that
the drug expires on the last day of February 1998. If the
text states use before February 1998 the drug expires on
the last day of January 1998

Q Most IP grade active ingredients be tested in accordance

with IP monograhs ?
Q Must manufacturers test each batch for all monograph
specifications ?

A The official speicifications are the minimum requirements of

a pharmaceutical compound. The in house specification are
tighter and more stringent than those laid that in the
Pharmacopeia.

Active ingredients must be tested for all specifications and

all batches must be characterised as per the specifications.

Q How is the moisture content of granules in a fluidised bed

drier measure ?

A Generally, in a fluidised bed drier, the drying is complete

when the outer temperature becomes equal to the inlet
temperature. By providing a digital temperature indicator,
and monitoring the outlet temperature the moisture content
of the material in a FBD may be determined. But, this is
not a pragmatic method and the best way to determine this
may be by the conventional IR balance method.

There is also a device developed in Germany whereby a

probe is placed inside the chamber of the FBD, which
measures the humidity and converts it to the moisture
content of the material.
Q What is the technical purpose a breakline on upper punch
and lower punch ?

A There is no technical purpose for providing break line on

upper punch and lower punch. Break line is provided only
for the convenience of taking half the dose if required.

Q What are the main raw materials used for manufacturing

membrane filters ?

A The most common filters in use are made of cellulose

esters like cellulose nitrate or acetate. Membranes are also
made from rayon, polycarbonate, nylon, polyvinylchloride,
polypropylene, teflon, and even gelatin.

Microporous membranes are made from a polymer solution

by a process of phase inversion. The factors involved are
temperature., water vapour inhibition and exchange of
solvents with non solvents. The pores are neither
cylindrical nor triangular but can be best described as
random polymeric froth consisting of 80% void volume.

Q What is the effect of steam sterilization on membrane

filters ? What happens at higher temperature ?

A Reverse phase membranes are the filters used almost

exclusively for sterilization by filtration. During casing
strain and molecular orientation are builts into the
membrane. The heating process may relieve the strain
Plasticization of the filter by heat occurs because the
viscocity of the filter is reduced at elevated temperature
enabling movement and adjustment of the polymer segments.
As a result, changes take place in the filter pores. Usually,
the pores shrink, and the bubble point of the filter rises. It
is also possible that some pores due to stress become
larger even while other are shrinking.

So it is a risk to repeatedly sterilize membrane filters.

In case of repeated sterilization, the bubble point test before
commencement of the filtreation proces is a must.

Q Why can’t we use lactose as diluent in isoniazid tablets ?

A Since lactose interacts with INH to form isonicotynyl

hydrazone, which is absorbed poorly from the GIT, it
should not be used in the formulation. More over, INH
 undergoes Mailard reaction with lactose and darkens the
colour of the tablet.

Q Does finished product ( FP ) testing provide us with an

unbiased view of the finished dosage form, or do we only
perform FP testing to satisfy the requirements of the
Regulatory Authorities ?

A Finished product testing in insolation does not provide an

unbaised view of the product. FP testing, done after the
process is complete, substantiates the results of the In
Process Control (IPC ) testing. To obtain an exact picture
both the IPC and the FP tests should be considered. This
would prove that the product conforms to its specifications.
FP testing cross checks the integrity of the product, and
therefore is more than a procedural formality.

If the testing proves conformance, the Regulatory

Authorities will be satisfied anyway.

Q Do IPC results carry more credibility than the results of

FP testing ?

A All organisations have established certain critical parameters

for their products, which are monitored by IPC. While IPC
documents and validates the process, FP testing is a natural
progression to conclusively proving the integrity of the
process. As FP re-establishes the claims of IPC, of the
product conforming to standards, it is equally important.

Q Should products tested always pass the FP specifications

before its release ? Can IPC data be taken into account if
the results obtained with EP are outside the specification ?
Should these results influence the decision to release the
product ?

A As stated earlier, IPC and EP testing should complement

each other. This proves that the process is under control.
In a fully validated process, if the IPC results meet the
requirements, the results of FP testing will meet the
specifications. When this is not so, resampling of the
finished product could be done. If the EP results are
outside specifications, the matter has to be completely
investigated, and the results documented. The final decision
 of release then rests with the Quality Control Department
based on their findings.

Q How do we set the IPC specifications and limits ?

A Normally, the pharmacopeial specifications are the basis on

which any process parameters are built. The IPC
specifications are more stringent than the pharmacopeial
limits, so as to make allowances for any possible variations
during the various stages of production. The FP limits too,
are equally if not more, stringent than the IPC limits.
These parameters are established during the validation
process.

Q Who should perform the IPCs ? Also, can IPC data

generated by production personnel be used for final
product release by the quality Control Department ?

Both the production department and the QC department

should perform the IPCs, independent of each other. The
data generated by the production department serves as their
guideline for controlling the manufacturing process.
Decisions taken by the Quality control must be based on
the compliance of results with specifications. IPC data
recorded by the production department can certainly be
taken into account by the QC if necessary. But, they make
the final decision on the release of the product.

Q Should the IPC and FP tests performed be derived entirely

as a result of validation studies ? If so, for established
processes, if the balance between IPC and EP testing is
modified, then should revalidation also be carried out ?

A IPC and FP tests are validated at the conception level of

any process. This is a joint exercise involving the R&D or
the Product Development, Production and Quality Assurance
personnel. Other departments like Engineering Supplier of
Raw Materials may also be involved. Revalidation becomes
necessary when there is any change in the process, change
in mixer capacity, modification of aseptic area.

It is recommended that the established process be validated

once in every six months.
Q What are the people / procedural factors which are
important to ensure successful application of an IPC / FP
testing system ?

Errors of any kind can cause have to a system in these

days of automation. People play and will continue to play
an important role in ensuring the success of any system.
The organisation should select and carefully train its
personnel to meet the demands of excellence required by
the Pharma Industry.

Operator training on any testing procedure needs a thorough

knowledge of the Standard Operating Procedures written for
that test to be executed. This step is probably the most
time consuming component of the training process.

Learning and applying a process not only requires physical

and intellectual abilities but also involves the development
and persistance of a correct mental attitude.

Q It is very difficult to handle materials which develop high

electrostatic charge, especially during dry granulation. What
can be done to neutralize ( partially or completely ), to
have a clean operation ?

A ESD ( electrostatic discharge ) or ZAP in Electronics and

Corona in the Pharma Industry, can trigger at even 1000
volts, white dry granulation reaches 35,000 volts at 15%
RH.

The solution lies in a proper salt pit earthing of the

granulator. Another solution is using only ionising process
air, after etablishing the electrostatic polarity of the
materials. Adding excipients that function as hygroscopic
cells ( any number of them both organic and inorganic )
which draw away, hold or dissipate the static charge could
also help.

Not any but all of these methods may be necessary.

Just a bit of triva Tajinder, check to cheek greeting can
tingle at 3000 Volts ?

Q Since the present US FDA regulations are objecting to the

fumigation of the area using formaldehyde and Potassium
Permanganate, due to post carcinogenic effects, is there any
effective method to fumigate the area ?
A Hydrogen peroxide Fumigation is the ideal method which
can be used.

Q Each LAF module comes equipped with a pressure gauge.

What does this pressure indicate ? How is it measured, i.e.
what are the two points considered to measure the
differential ? Is the pressure measured in terms of water is
necessary ? What are the limits of pressure beyond which
one should get the module checked ?

A Each LAF unit is equipped with a manometer or a

magnabelic gauge to indicate the plennum pressure with
respect to ambient. The indicated pressure itself is the
differental pressure differential with respect to ambient.

The plenum presure may vary slightly from equipment to

equipment. This is due to the density of the HEPA media.

When equipment is installed and demonstrated, a note

should be made of the initial indicated pressure, as this
becomes the reference base for future.

When a prefilter gets choked up, the intake air quantity is

reduced, thus building lesser plenum pressure with respect
to initially noted manometer ( or gauge ) reading. This is an
indicator calling for action “ Pre Filter Needs Cleaning “

When the HEPA filter gets choked up, the air flow is
reduced. The plenum pressure naturally increases beyond the
initially noted reading. Normally all HEPA filter have a
longevity beyond two years, subject to disciplines
maintained and the plenum could go upto 17 to 18 mm,
beyond the previously noted pressure reading. When the
pressure goes up, and velocity reduced below 0.4 M/sec.
HEPA filter have to be changed.

A check should be carried out once in every six months to

assure the quality air is maintained continuously.
Manometer pressure reading is alwa in millimeters of water
not mercury. So is the case with magnethlic gauges.

Q What is the best SOP for checking the accuracy of marking

line, such as 2.5 ml, 5 ml in deep coloured plastic
measuring glass, provided with liquid oral bottles ?
A Purified water is added upto the marking line using a
calibrated pipette and the volume delivered is measured.
The same procedure is followed even for coloured / opaque
measuring cups. It is generally advisable to provide
transparent measuring cups for the convenience of the
consumer.

General information section ( 1221 ) of USP, under the title

teaspoon, specifies that the volume error incurred in
measuring liquids using such devices, should be, not greater
than 10% of the indicated amount.

Q What is the requirement for intensity of light required for

carrying out various operations in the manufacturing of
tablets and parenterals ?

A The intensity of light required in the manufacturing area

for carrying out various processes is 300 to 400 lux. For
visual inspection, the intensity required is 160 feet candle
at the working level.

Q How can we reuse the enteric coated tablets rejection ?

A GMP requirements permit reuse only if the quality of the

final product is not affected and additional testing of
finished product, into which a recovered product has been
incorporated. Is carried out, if required.

Effect on the product attributes. On introduction of

polymers used for enteric coating in the core tablets, is
less predicatable. It may affect the physical parameters as
well as availability of the drug adversely. Therefore, reuse
of enteric coated tablets, either by intorudction into a
subsequent batch or, by processing the pooled residue as a
separate batch, is not advisable.

Q What are the sanitizing solutions to be used, with rotation

and frequency for.

a Cleaning of floors
b Cleaning of equipment
c Epoxy coated walls and glass panels

Unique Pharmaceutical Labs.

A For sanitization of floors, walls, and glass panels, Lysol
(Cresol ), Dettol 5% ( Chloroxylenol ) Savlon 2%
(Chlorohexidine Gluconate and Cetrimide ), are
recommended. The disinfectants are used in rotation and
should be changed on a weekly basis.

For cleaning of equipment, use of 0.1% sodium lauryl

sulphate solution and for sanitization of equipment use of
filtered steam is recommended. For drying of washed
equipment, infra-red driers are being used currently, which
avoids the use of mops.

Effective cleaning is 90% of the overall sanitizing job and

application of sanitisers is the remaining 10%. Therefore
sanitization responsibilities should be performed by trained
and reliable people.

Q What is the best SOP for validation of ampoule washing

machine.

A Validation of a machine will depend upon the type of

washing process employed, such as jet washing, ultrasonic
cleaning before jet washing etc. The operating variables
generally change with the type of machine used.

Control of these variables during the washing process

should be established before checking the cleaning
performance. Some of the important variables and necessary
verifications to be carried out are listed below :

* All the measuring devices should be calibrated.

* Adequacy of the air and water pressure should be

confirmed.

* Cleanliness and proper mounting of air and water filters

should be confirmed .

* It should be ensured that all the washing needles are open

and the required quantity of water is delivered in each
washing cycle.

Effectiveness of cleaning can then be checked on washed

ampoules as described below :
* The washed ampoules of a wash cycle are collected under
a laminar air flow work station.

* The ampoules are filter with distilled water, filtered

through 0.2 micron membrane filter and the open ends are
closed with aluminium foil.

* These ampoules are visually examined for presence of

particulate matter.

The washing process can be continued if the percentage

rejects are found to be within acceptable limits.

Q What are the methods of sampling in the course of

evaluation of cleaning validation ?

A There are two general types of sampling that are

acceptable. The best method is direct sampling of the
surface of the equipment. The second method is the use of
rinse solution.

Direct surface sampling :

The first step is determined by the type of sampling

material used and its impact on the test data, since the
sampling materials may interfere with the test. It is very
important, early in the validation program, to ensure that
the sampling medium and the solvent are satisfactory and
can be readily used.

Advantages of direct sampling are that, the areas hardest to

clean and which are reasonably accessible can be evaluated,
thus establishing a level of contamination or residue per
given surface area. Additionally, residues that are dired out
or are insoluble, can be sampled by physical removal.

Rinse sampling :

The advantage of rinse sampling is that a larger surface

area may be sampled and systems that cannot be routinely
disassembled can be sampled and evaluated.

Disadvantages of the method are that the residue or

contaminant may not be soluble or may be physically
occluded in the equipment.
 Routine production In process Control.

Monitoring and indirect testing, such as conductivity testing

may be of some value for routine monitoring once a
cleaning process has been validated. This could particularly
be true for the bulk drug substance manufacturer where
reactors and centrifuges and piping between large equipment
can be sampled only using rinse solution samples. During
validation, the firm should document that, testing the
uncleaned equipment gives not acceptable result for the
indirect test.

Q Is there any other method of depyrogenation other than dry

heat ?

A Apart from dry heat sterilization, there are two method of

depyrogenation i.e. by endotoxin removed and by endotoxin
inactivation.

Endotoxin removal can be achieved by the following

methods :

Rinsing

This is the oldest and simplest method of removal of

pyrogens from solid surfaces. These are rinsed with non
pyrogenic solvents, usually sterile water for Injection. Low
levels of surface endotoxin contamination can be effectively
removed from glassware, device components and stoppers.

Rinse water can be monitored throughout the process with

LAL to validate endotoxin removal.

Distillation

This method is used to remove pyrogen from water.

Water is forced to undergo two phase changes, from liquid
to vapour and from vapour to liquid. Because the
lipopolysaccharide is a large molecule, it can not boil as
rapidly as water and is left behind. Those molecules
entrained in water droplets carried in the stream are
dropped back by gravity due to their high molecular
weight.

* Ultra Filtration
Ultra filtration membranes are rated on the basis of
molecular weight exclusion limits and their effectiveness as
depyrogenating filters is due to their action as size
discriminating screen. Thus, endotoxin that exceed the
molecular weight exclusion limit of a given membrane are
retained on the surface of the membrane.

The basic sub-unit size of a lipopolysaccharide is 10,000 to

20,000. It can therefore be effectively removed from
solutions by a 10,000 molecular weight ultra filter.

* Reverse Osmosis

Reverse osmosis membranes consist of cellulose acetate or

polyamide materials with pores small enough to exclude
ions. Conventional reverse osmosis membranes ( normally
rated pore size 10 A ) remove endotoxin by simple size
exclusion. The pores in the membrane are far too small to
pass the pyrogens.

* Activated Carbon

Depyrognation of solutions can be done based on the

physical adsorption of endotoxin to charcoal particularly
activated charcoal. Charcoal is added to the solution and
agitated and the carbon is removed by filtration. This is
widely used in pharmaceuticals. But carbon has a tendency
of absorbing the active ingredients in the solution since it
has an affinity for high molecular weight substances.

* Electrostatic attraction

This is achieved by using depth filtration, using asbestos

which adsorbs the endotoxins. Endotoxin Inactivation can be
done by the following methods :

* Acid base hydrolysis

* Oxidation
* Alkylation
* Moist heat treatment

Normal autoclaving is not effective for depyrogenation, only

long drastic heating would destory pyrogens. Example
autoclaving at 20 psi for 5 hrs. At pH 8.2 or 20 psi for
2 hrs. At pH 3.8.
 * Ionizing Radiation
using 60 C.

* Using Polymyxin B
These are the different methods of depyrogenation which
can be used based on suitability. Each process should be
validated before use.

Q We are calibrating HPLC equipment. I.e. pump ( flow check )

detector, injector / auto injector and system suitability tests
as per USP XXIII It is recommended that this calibration
be done once a month. We are ensuring that the
percentage relative standard deviation of six replicate
injections of standard preparation is below 1.5%

For day to dry routine product release, is it necessary to

take six replicate injections of standard preparation all the
time ? Or is monthly calibration data sufficient ?

A The USP XXIII requires that unless otherwise specified in

the individual monograph, five replicate injections of the
analyte are required for calculation of the RSD, if the
RSD is 2% or less, and six replicate injections, if the
RSD is greater than 2%. Hence, if one wants to comply
with the USP in totally it will be essential to carry out
the exercise on a day to day basis. However, if the
wording old preparations in the question means established
products for which the HPLC method in use has been fully
validated and adequately documented, then it is possible to
carry out triplicate or duplicate injections for day to day
analysis and follow the USP protocol at defined intervals
of say 3 months, as a part of revalidation exercise. This
deviation from the the USP must be covered by an SOP.

Q What is the calibration frequency for mercury in glass

thermometers ? I understand that glass thermometer
calibration is valid for a life time. Is it so ?

A Mercury in glass thermometers can remain within calibration

for several years, provided the same is stored and handled
in a controlled manner. However, unless a thermometer is
identified as a master calibrated thermometer, and stored
and handled in a controlled manner as defined in an SOP,
thermometers must be calibrated annually as part of GLP.
Q How are humidity control ovens calibrated for both
humidity as well as temperature ?
A Humidity, control ovens can be calibrated for humidity
using a calibrated wet and dry bulb thermometer, or by
using primary standard chemicals, which can give
established humidity in saturated solutions in contact with a
solid. For temperature calibration, use any calibrated
thermometers.

These questions have been answered by Mr. R.

Raghunandan. Corporate Quality Manager, Glaxo India Lts.

Q What kind of regulatory clearances does one need to have

for exporting Ayurvedic formulation ?

A If an Ayurvedic formulation is permitted to be manufactured

and marketed in India, we would not require any other
clearances from the Drug Control Administration for
exporting the Ayrvedic formulations. However, there is a
ban on the export of certain plant materials, either in the
crude form, their extracts or formulations.

Q Should silicon emulsion used to coat vials and stoppers be

filtered ? What is the most effective method for filtering
silicon emulsion ?

A The silicon emulsion used for coating should definitely be

filteed. This is done to free the emulsion from particulate
matter which might settle on the lining. Any particulate
matter settling on the vial would results in poor coating.

The best method is to filter through a 0.3 micron cellulose

membrane filter.

Q What is the normal sampling time and sampling volume

using an air sampler. What would be the ideal sampling
procedure ? What should be the frequency of sampling ?

A The normal sampling volume using an air sampler, would

be 3000 litres at a time, at a particular place. It is
essential to make same that the corners of the room are
definitely included in the sampling. Other than that, two or
three places, in the room should be sampled. The number,
position and sample volumes should reflect the standard of
control required, the level of occupancy and activity.
 Monitoring for class V1 and class hundred should be
carried out as close to the critical zone as possible. This
may require a considerable developmental input to establish
a practical method.

The frequency of sampling is determined by in house

practice, based on procedures that have been validated.
However, the following is the frequency as indicated in
contamination control practice.

Class Frequency

V1 Daily or batchwise
V2 Daily
V3 Daily
V4 / V5 Weekly
100 Daily
1,000 Daily
10,000 Weekly

Q What are the optimum storage conditions prescribed for

rubber stoppers ? How should they be handled ?

A Rubber stoppers, before sterilization, should be stored in

HDPE bags, in loosely packed numbers, as otherwise they
would tend to stick to each other. These bags are stored
at room temperature ( less than 35 C ) at a relative
humidity of about 45% The stoppers should be protected
from dust as the dust tends to stick to the stoppers and
their removal is very difficult. They should also be
protected from grease and oil as rubber stoppers absorb oil
and swell up.

The stoppers are sterilized immersed in water for injection

along with a preservative double the concentration of that
used in the final product. The level of water should be
enough to compensate for any loss during the sterilization
process.

If it is required that the stoppers should be dried, the

drying should be done in a hot air oven fitted with HEPA
filters, at temperatures less than 100 C Repeated
sterilization of the stoppers should be avoided. If the
stopper are to be stored, it is better that they be stored in
a dry condition.