“ROLE OF INSTRUMENTATION IN

QUALITY CONTROL IN PHARMACEUTICAL

INDUSTRIES”

A dissertation submitted to

DR.REDDYS LABORATORIES LTD. BANASTHALI

VIDHYAPITH
FTO UNIT –VI, Baddi BANASTHALI,
RAJASTHAN

IN THE PARTIAL FULFILLMENT FOR THE AWARD OF

MASTERS OF SCIENCE
IN
PHARMACEUTICAL CHEMISTRY
2008-2009

BY
SONALI SHARMA

Guide:
Mr. Rajesh M Sharma Dr.. Sarvesh Paliwal
H.O.D., Associate Professor,
Quality control, Deptt. Of Chemistry
Dr.Reddy’s lab. Ltd. Pharmaceutical chemistry,
FTO Unit-6, Baddi, Banasthali vidhyapith,
 TABLE OF CONTENTS
• CERTIFICATE

• ACKNOWLEDGEMENT

• NTRODUCTION
 Company profile

• QUALITY CONTROL
 Instrumentation

 Karl fischer moisture content detector

 Polarimeter

 Ultra-violet and visible

spectrophotometry

 Gas chromatography

 High performance liquid

chromatography
• QUALITY ASSURANCE
 CERTIFICATE
This is to certify that Miss Sonali Sharma, final year student of M. Sc.
Pharmaceutical Chemistry, Banasthali Vidyapith, Banasthali, Rajasthan,
did her project work on “Role of instrumentation in quality control in
pharmaceutical industries” for the partial fulfillment of the award of
Master’s Degree. She worked from 12th January to 21st April, 2007 under
my supervision in the Dr.Reddy’s Laboratories Ltd. FTO Unit-6, Khol,
Baddi - Himachal Pradesh.

The data contained in her report has resulted from the work that she
carried out during the tenure of her training program. This work was
originally carried out in my laboratory by various workers.

I wish her all the success in her life.

Mr. Rajesh M Sharma

Head
Quality Control
Dr. Reddy’s Laboratories Ltd.
FTO Unit – VI,
Khol village, Baddi (HP)
 DECLARATION

I hereby declare that the dissertation entitled “Role of instrumentation

in quality control in pharmaceutical industries” submitted to
Banasthali Vidyapith, Rajasthan and Dr.Reddy’s laboratories ltd. khol
baddi, in partial fulfillment for the award of M.Sc. Pharmaceutical
Chemistry is a record of research work done by me under supervision and
able guidance of Mr. Rajesh sharma, H.O.D quality control.

I also declare that nothing in part or full has been submitted for the award
for any other degree, diploma, or fellowship to any other university or
institute.

Sonali Sharma
 ACKNOWLEDGEMENT

A journey is easier when you travel together. This project report is the
result of five months of work whereby, I accompanied and supported by
many people and its easant time to express my gratitude for all of them.

Fore most, I would like to thank God for His grace and blessings.

The first person I express to my heartiest thanks to Mr. Rajesh M

Sharma, Head, quality control department , for providing me excellent
laboratory facilities, valuable guidance, untiring help during my project
work and constant encouragement during the whole period of my project
work

I wish to express to my sincere gratitude to Mr. Surinder Pal Singh, HR,

Dr.Reddy’s lab, FTO Unit-6, baddi for giving me an opportunity to carry
out my project work in such a prestigious institute.

I am grateful To Mr. Vikrant Singh Parmar, Mr. Ashwin Deep Mishra,

Mr. Ashwani Mahajan, Mr. Bipan Singh Katoch who have provided
support at each and every step of my work.

This acknowledgement seems to be incomplete without the name of Miss.

Jyotsna chauhan, Miss. Priyanka mishra, Miss. Babita. Whose
consistent help has only leaded me to the completion of this project work.
I am also thankful to Mr. Dharmender verma for enriching me during
course of investigations.
I owe my gratitude to Prof. D. Kishore, H.O.D., Deptt. Of Chemistry and
Pharmaceutical Chemistry, Banasthali Vidyapith, Rajasthan and Mr.
Sarvesh K. Paliwal for their kind support throughout the course.

The chain of gratitude would be incomplete, if I forget the name of my

friend Miss. Vaishali jamwal with her I enjoyed a lot at Dr.Reddy’s.

Finally, I express my gratitude to my family members for their blessings

and constant support. Words with me are insufficient to express the
feelings of my heart to acknowledge them for educating me and keeping
me in all comfort without which this work would not have seen the light
of the day at all.

Sonali sharma
 COMPANY’S PROFILE

DR.REDDY’S LABORATORIES PROFILE

Dr. Reddy’s aim is to help people lead healthier lives through two parallel
objectives:
Making medicine affordable and accessible in all parts of the world so
that many people as possible benefit from them.

And discovering and developing and commercializing innovative

treatment options that satisfy unmet medical needs.
With headquartered in India, it is a global pharmaceutical company with
a presence in more than 100 countries. Dr. Reddy’s is the first
pharmaceutical company in Asia outside of Japan to be listed on the
NYSE.

Its strong portfolio of businesses, geographies and products gives an edge

in an increasingly competitive global market and allows it to provide
affordable medication to the people across the world, regardless of
geographic and socio-economic barriers. Dr. Reddy’s is a global,
vertically integrated pharmaceutical company with a presence across the
value chain, producing and delivering safe, innovative and high quality
finished dosage forms, active pharmaceutical ingredients and biological
products.

It conducts NCE drug discovery research in the areas of metabolic

disorders and cardiovascular indications as research facilities in Atlanta
(USA) and Hyderabad (India). Through its custom pharmaceutical
services business unit, it provides drug substance and drug product
development and manufacturing services on a proprietary basis.

PHARMACEUTICAL CHEMCALS
Active pharmaceutical ingredient: its capabilities span 24
major chemistries including stereo selective synthesis,
cryogenics, hydrogenations and cyanations. Its strong IP and
analytical skills are evident in 84 US DMFs it has filed, the
highest in India and the second highest in the world. Its
manufacturing facilities are capable of supporting the product
development effort through concurrent scale-up and piloting of
feasible route as they are developed by the R&D teams. State-
of-the-art equipment and instruments give it’s the edge to
compete globally. Its operations are fully integrated through
supply chain and ERP systems (SAP R/3), which enable
seamless response to customers, all the while keeping the
environment around plants clean, green and safe.

Custom pharmaceutical services: In an industry cluttered with

manufacturers, CPS stands out because of its understanding of
pharmaceutical business and the associated expertise needed. Rather than
just being a chemical provider, CPS offers a service mix covering the
entire pharmaceutical value chain. It execute cost-effective and time-
bound projects for its customers, and provide them with cGMP-complaint
products manufactured in FDA-inspected, ISO-certified facilities.
A team of experienced project managers ensures smooth progress if
projects from initiation to closure in order to avoid any cost and time
overruns.

GENERICS

GENERIC FORMULATIONS: Geographic diversifications, cost

containment, strengthening its product portfolio and building scale –
Dr.Reddy’s is strong in all these aspects in the generic space. It is now
the fourth largest in Germany after the acquisition of betapharm, and is
constantly looking for opportunities to maximize the potential of its
current and future portfolio in different territories across the US and EU.
It has the necessary expertise of customer-specific packaging, compliance
packaging and anti-counterfeit packaging. In fact Dr.Reddy,s has won
several awards globally fir its packaging efforts, including the Asia star,
Ameristar and World star awards.

Branded formulations: Dr.Reddy’s brand are todays recognized and

trusted across several continents. Brands like Omez (omperazole), Nise
(nimesulide), Stamlo (Amlodipine), Ciprolet (ciprofloxacin), Enam
(Enalapril) and Ketorol (Ketorolac) are leaders in their category in several
countries, with many of them being used by more patients than use the
innovators products. Over 1.5 million patients across the world take
‘omez’ for their acid peptic disorders every single day! Entrepreneurship,
coupled with the will to make a difference drives our 2,000 strong field to
force to reach out to over 210,000 doctors and 115,000 pharmacies in
more than 40 countries across the world.

INNOVATION

DISCOVERY REASEARCH: It has put in place a state-of-the-art,

fully integrated discovery infrastructure to strengthen its effort to
discover and develop therapeutically useful new chemical entities (NCEs)
and market them globally. Its two discovery research centers one in
Atlanta. USA and other in Hyderabad, India have more than 300 hundred
scientists actively involved in a number of drug discovery and clinical
developments programs.

Specialty pharmaceuticals: Its specialty pharmaceuticals business

deals with assets like acquired proprietary technologies, internally
developed proprietary drug- delivery platforms, and current internal
compounds under pre-clinical and clinical development. Its initial global
therapeutic area focus is on dermatology and oncology, two therapeutic
areas that best leverage its internal assets. A key component of the
strategy in this area is strong, targeted business development effort to
accelerate market entry.

Biopharmaceuticals: Its biologics development center span an area

of 36,000 sq. ft., with development and manufacturing suites foe both
E.coli and mammalian cell culture. It caters to the highest development
standards of cGMP, GLP and applicable levels of bio-safety.
Grafeel (Filgrastim), its first biologics product to enter the market, enjoys
a market share of almost 50% in India and has able to reach many more
patients than the innovators product due to its affordability. Its second
product Reditux (Rituximab) is the first biosimilar monoclonal antibody
to be developed and launched anywhere in the world.

PHARMA SERVICES AND ACTIVE INGREDIENTS

The global API business of Dr. Reddy’s serves generics and innovator
companies through its API (Active pharmaceutical ingredients, also
called ‘bulk actives’) and CPS (Custom pharmaceutical services)
businesses respectively.

Dr. Reddy’s began APIs operations in 1984 and started with a single drug
in a 60-tonn facility near Hyderabad, in India. In 1986, the first
consignment of that drug, methyldopa, was shipped to West Germany. Its
emphasis on high quality and R&D led to USFDA inspection of our
manufacturing facilities before long. Since then, Dr. Reddy’s has come a
long way and today has a wide portfolio of APIs. Its core strength in APIs
has continued to increase significantly over the years. It has the largest
number of US DMF submissions from India and are among the top five
API players globally. Its global API business offers over 100 molecules
to customers across the world.

Its CPS business, formed in 2001, serves more than five big
pharmaceutical companies and over 25 emerging pharma companies
today. The CPS business got a boost in 2005 with the acquisition of
Roche’s API manufacturing unit in Mexico. This facility added a niche
capability of steroid API manufacture to DRL’s capability portfolio.
Today, Dr. Reddy’s is the largest CPS player from India. Its CPS
business provide a unique opportunity for innovator companies
worldwide to make use of its technical expertise, world-class
infrastructure, and flexibility to bring their medicines to the market
quickly and economically. Dr. Reddy’s has emerged as a trusted supplier
of value added advanced intermediates and APIs to generic as well as
innovator companies.

The discovery and development of new active ingredients for a medicine

is a long and expensive process which is why a patent protects a new
active ingredients for a certain a time. When the patent expires, other
pharmaceuticals companies market medicine with the same active
ingredient
 KARL
FISCHER MOISTURE
CONTENT DETECTOR
 POLAR
IMETER

 ULTRA
-VIOLET AND VISIBLE
SPECTROPHOTOMETRY

 GAS
CHROMATOGRAPHY

 HIGH
PERFORMANCE LIQUID
CHROMATOGRAPHY
 QUALITY CONTROL

Quality is important in every product or service but it is

vital in medicine as it involves life. Unlike ordinary consumer
goods there can be and there is no 'Second' quality in drugs.

Quality control is a concept, which strives to produce a perfect

product by series of measures designed to prevent and estimate
errors of different stages of production. As a mater of fact it is
built in from the time of inception of the thought to make a
product, to the time, it is finally made and sent out with an ok
quality report. In popular practices the quality of medicines or
pharmaceutical product is assured through quality control. It is,
therefore, essential that quality assurance department must adopt
"Good Laboratory Practice" to ensure reliability and accuracy of
results given out by them.
The assurance of the quality and the reliability of
pharmaceuticals, together with their careful control are out
moral obligations arising from the humanism towards the sick
human beings. Consequently, the manufacture and the control of
the drugs are very responsible tasks and they need substantial
knowledge of the science.
The decisions to release or reject a product are bared upon one
or two types of control actions or combination of both.
Providing sample analytical procedures for complex
formulations is a matter of foremost importance.

QUALITY CONTROL:-

QC department is just like the backbone of the company because

the quality and the feasibility to stand and the market are
checked here. Products manufactured are checked for various
parameters required to be fulfilled before they are dispatched to
the market.
Quality control department consists of the analysis results from
three different labs.

Instrumentation Lab :- It is also called as the quality insurance

lab and it mainly deals with the chromatographic techniques
lime High Performance Liquid Chromatography (HPLC) and
Gas Chromatography to mark the potency and the efficiency of
the drug, This lab deals with the following experiments:-

 Assays: - This is mainly done to check for the potency of

the drug produced.
 Residual Substrate Estimation: - This test is performed to
check for the residual substances present in the resultant
drug.
 Reaction Monitoring :- This test is of utmost importance as
through this test we continuously keep a check on the
reaction carried out in the reactor by analyzing the
intermediated formed at different step

QUALITY CONTROL AND ITS RESPONSIBILITIES

Quality control is the part of good manufacturing practices

concerned with sampling specifications & testing & with the
organization, documentation and release procedures which
ensure that the necessary and relevant tests are actually carried
out and that materials are not released for user nor products for
sale or supply until their quality has been judged to be
satisfactory. Quality control is not confined to laboratory
operations but must be involved to laboratory concerning the
quality of the product.
 Each holder of the manufacturing authorization should
have a quality control department. The independence of quality
control from production is considered fundamental. The quality
control department should be independent than the other.
The basic requirements for quality control are as follows:

1. Adequate facilities, trained personnel and approved

procedure must be available for sampling inspecting and
testing starting materials, packing materials and
intermediate, bulk and finished products and where
appropriate for monitoring environmental conditions for
GMP purpose.
2. Sample of starting materials, packing materials,
intermediate products bulk products and finished
products must be taken by methods and personal approved
of b the quality control department.
3. Test methods must be validated.
4. Record must be made demonstrating that all the required
sampling, inspecting and testing procedures have actually
been carried out and that any deviations have been fully
recorded and investigated.

The quality control department as a whole will also have

other duties, such as to establish validate and implement are
quality control procedures, to evaluate, maintain and store the
reference standards for substances to ensures the correct labeling
of containers of materials & products to ensure that the stability
of the active pharmaceutical ingredients and products is
monitored, to participate in the investigation product and to
participate in the investigation products and to participate in
environmental monitoring all there operations should be carried
out in accordance with written procedures and where necessary,
recorded.

QUALITY SYSTEM
The basic elements of quality and systemic action. The
quality system involves all phase from initial identification to
final satisfaction of requirements and customers expect action.
The first thing that a manufacturer will like to know is
what product should be manufactured. The next steps follow as
under:
Procurement of raw materials
↓
Process planning
↓
Production
↓
Inspection & test
↓
Packing
↓
Storage
↓
Sales & Distribution

KARL FISCHER MOISTURE CONTENT

DETECTOR

For the determination of small amounts of water, "Karl

Fischer" in (1935) proposed a reagent, is known of Karl Fischer
reagent.
Principle
In this method, the action of sulphur dioxide upon a
solution of Iodine in a mixture of anhydrous pyridine and
anhydrous methanol. Water reacts with this reagent in a two
stage process in which one molecule of iodine disappears for
each molecule of water.

CH3OH + SO2 + RN → RNH – SO3R

R5HNSO3R + 2RN + I2 + H2O → (RNH) SO4R + 2 (RNH) I

The end point of the reaction is conveniently determined

electrometrically I2 generate electrometrically from I-.

When iodine comes in contact with water the reaction as:

I2 + SO2 + H2O → HI + H2SO4-

The amount of water in the sample is calculated by measuring

the current needed for electrochemical generation of iodine from
I -.

2I- → I2 + 2e-
Apparatus
A titration vessel of about 60ml capacity is fitted with two
platinum electrodes about 0.05 sq. cm in area and about 2.5cm
apart, a nitrogen inlet tube, a stopper which accommodates the
burette tip and vent tube protected by a suitable desiccant such
as phosphorous pent oxide or silica gel. The substance being
examined is introduced through an Intel which can be closed by
a ground stopper. Stripping is done by magnetically. The air in
the entire system should be kept dry during the titration.
The voltage across the electrodes may be obtained from a
1.5volt dry cell and a variable resistance of about 200ohms. The
resistance is adjusted so that an initial current passes through the
electrodes connected in series to an ammeter. On adding the
reagent the needle of the ammeter shows a deflection but return
immediately to its starting position. At the end point of the
titration a slight excess of the reagent produces a deflection
which persists for not less than half a minute.

Procedure
The determination of small amount of moisture content involves
three steps.
1. Neutralization of methanol
a) Fill the titration of vessel with methanol in order to
dip the Pt. electrode in it.
 b) Start the titration in order to neutralize the methanol.

2. K.F. Factor
a) Add about 150 to 200ml of purify water in titration.
b) Start the titration and not the volume of K.F. reagent
used.

K.F.Factor = Wt. of purified water

Volume of K.F.Reagent used

Limit = 5-7mg/ml.

Units = mg/ml

3. Moisture Content
a) Wt. amount of sample according to STP add it in
titration vessel.
b) Stir it for one minute
c) Titrate it with K.F. reagent, note the volume of K.F.
Reagent.

V × F × 100
% of moisture content = ----------------
W × 1000
V = Vol. of K.F.R. used with sample.
F = water equivalence factor
W = weight of sample.

Calibration
Standardization of KF Reagent
1) Remove all the liquid from KF apparatus vessel, rinse it
with methanol twice and transfer about 30 ml methanol
into vessel.
2) Neutralization to end point with KF reagent.
3) Weight accurately about 150mg of disodium tartrate
transfer it into KF vessel, wait for about 10-15 seconds.
Start addition of KF reagent record the volume of KF
reagent consumed.

Calculate the KF factor by the formula

W × 1000 × 15.66
F is mg/ml = --------------------------
V × 100

W = weight of disodium tartrate in g

V = Volume of KF reagent consumed in ml.
Note:
Disodium tartrate can be replaced with water for
standardization with the change in step no. 3 ice weight
accurately about 30-6 mg of water, transfer it into KF vessel,
start addition of KF reagent record the vol. of KF reagent
consumed and calculate the factor by formula.

W × 1000
F mg/ml = --------------
V
W = weight of water in g
V = vol. of KF reagent in ml.

POLARIMETER

Many pharmaceutical substance are optically active i.e.,

they rotate an incident plane of polarized light so that the
transmitted light emerges at a measurable angle to the plane of
the incident light. This property is characteristic of some crystals
and of many pharmaceutical liquids or solutions of solids. The
property is the result of presence of one or more asymmetric
centers usually a carbon atom with four different asymmetric
centers. The only convenient means for distinguishing optically
active isomers from each other is polarimeter.
Substances that show optical rotation are optically active.
Those that rotate light in a clockwise direction as viewed
towards the light source are dextrorotatory. Those that rotate
light in the opposite direction are called laevorotatory.
The general equation used in polarimeter is:
λ
[D] = 100 α
I. C
[D] = specific rotation at wavelength λ
t = temperature
a = observed rotation in degrees
I = path length in decimeter
C = concentration of the substance

Polarimeter was performed using an instrument where the

extent of the optical rotation is estimated by visual matching of
the intensity of split fields. For this reason the D-line of the
sodium lamp at the visible wavelength of 589nm was most often
employed.

Instrument:
Optical System
Provide basic optical rotation information. It contains a
polarizer section, sample chamber and analyzer section.
Power Module
It transforms the 110 or 220v a/c line power into the AC
and DC voltage for the system.

Procedure:
 Take a polarimeter tube that has been selected, cleaned
and tested.
 Fill the tube with the solvent used to dissolve the optically
active sample and place in the sample chamber.
 Press zero reset button.
 Remove the solvent from the tube and rinse at least three
times with sample solution.
 Fill with sample solution and place in sample chamber.
 After achieving polarimetric balance, record the display
reading.

Applications The physicochemical properties of non-super

imposable chiral substances rotating plane polarized light in
opposite directions to the same extent, enantiomers are identical
except for this property and in their reactions with other chiral
substances. Enantiomers often exhibit different pharmacological
and toxicological properties, owing to the fact that biological
receptors and enzymes themselves are chiral. Many article from
nature source, such as amino acids, proteins, alkaloids,
antibiotics, glycosides, and sugars, exist as chiral compounds.
Synthesis of such compounds from non-chiral materials results
in equal no. of enantiomers, recreates. Recemates have net null
optical rotation and their physical properties may differ from
those of the componentanatomies.

ULTRA VOILET AND VISIBLE

SPECTROPHOTOMETRY

When radiation is passed through a layer of a solution

containing an absorbing substance, part of the radiation is
absorbed; the intensity of the radiation emerging from the
solution is less than the intensity of radiation entering it. The
magnitude of the absorption is expressed in terms of the
absorbance, A, defined by the expression.
A = log10 (I10/I)
Where I0 is the intensity of the radiation passing into the
absorbing layer and I is the intensity of the radiation passing out
of it. The absorbance depends on the concentration of the
absorbing substance in the solution and the thickness of the
absorbing layer taken for measurement. For convenience of
reference and for ease in calculations, the absorbance of a 1-cm
layer of a 1% w/v solution is adopted for several substances and
evaluated by the expression.
A (1%, 1-cm) = A/C l
Where c is the concentration of the absorbing substance
expressed as percentage w/v and I is the thickness of the
absorbing layer in cm. The value of A (1%, 1-cm) at a particular
wavelength in a given solvent is a property of the absorbing
substance.

Apparatus
A ultra-violet and visible spectrophotomer, suitable for
measuring in the ultra-violet and visible range of the spectrum
consist of an optical system capable of producing
monochromatic light in the range 200 to 800mm and a device
suitable for measuring the absorbance.
 Two empty cells used for the solution being examined and
the reference liquid must have the same spectral characteristic.
Where double beam recording instruments are used, the solvent
cell is placed in the reference beam.

Control of Wavelength
Verify the wavelength scale using the absorption maxima
of holmium per chlorate solution, the line of hydrogen or
deuterium discharge lamp or the lines of a mercury vapour are
shown below:
The permitted tolerance is ± 1 nm for the range 200 to
400nm and ± 3 nm for the range 400 to 600nm.

241.15nm (Ho) 404.66 nm (Hg)

253.70nm (Hg) 535.83 nm (Hg)
287.15 nm (Ho) 486.00 nm (Dβ )
302.25nm (Hg) 486.10 nm (Hβ )
313.16 nm (Hg) 536.30 nm (Ho)
334.15 nm (Hg) 546.07 nm (Hg)
361.50 nm (Hg) 576.96 nm (Hg)
365.48 nm (Hg) 579.07 nm (Hg)

Control of Absorbances

Check the absorbance using potassium dichromate

solution UV at the wavelength indicated in Table1, which gives
for each wavelengths the exact value of a (1%, 1cm) and the
permitted limits.
 Table – 1
Wavelength (nm) A (1%, 1cm) Maximum
tolerance
235 124.5 122.9 to 126.2
257 144.0 142.4 to 145.7
313 48.6 47.0 to 50.3
350 106.6 104.9 to 108.2

Limit of stray light

Stray light may be detected at a given wavelength with
suitable filters or solutions; for example, the absorbance of a
1.2% w/v solution of potassium chloride at a path length of 1cm
should be greater then 2.0 at about 200nm when compared with
water as reference liquid.

Resolution power
When prescribed in a monograph, record the spectrum of
a 0.02% v/v solution of toluene in hexane UV. The ratio of the
absorbance at the maximum at about 269nm to then at the
minimum at about 266nm is not less than 1.5 unless otherwise
specified in the monograph.
Spectral slit width
When measuring when measuring the absorbance at
absorption maximum the spectral slit width must be small
compared with the half-width of the absorption band otherwise
erroneously low absorbance will be measured. Particular care is
needed for certain substances and the instrumental slit width
used should always be such that further reduction does not result
in an increased absorbance reading.

Cells
The absorbance of the cells intended to contain the
solution to be examined and the reference liquid, when filled
with the same solvent, should be identical. If this is not the case,
an appropriate correction must be applied. The tolerance on the
path length of the cells used is ± 0.005cm. Cells should be
cleaned and handled with great care.

Solvents
In measuring the absorbance of a solution at a given
wavelength, the absorbance of the reference cell and its contents
should not exceed 0.4 and should preferably be less than 0.2
when measured with reference to air at the same wavelength.
The solvent in the reference cell should be of the same lot as
that used to prepare the solution and must be free from fluoresce
at the wavelength of measurement. Ethanol (95%), ethanol,
methanol and cyclohexane, used as solvents, should have an
absorbance, measured in a 1-cm cell at about 240nm with
reference to water, not exceeding 0.10.

Determination of absorbance
Unless otherwise directed measure the absorbance at the
prescribed wavelength using a path length of 1cm at 240 to 260.
If necessary, this path length may be varied provided that
compliance with Beer's law has been shown over the range in
question. Unless otherwise prescribed, carry out the
measurements with reference to the solvent used to prepare the
solution being examined. In certain cases measurement are
carried out with reference to a mixture of reagents, detail of
which are given in the individual monograph.

A statement in an assay or test of the wavelength at which

maximum absorption occurs implies that the maximum occurs
either precisely at or within ± 2nm of the given wavelength.
Likewise a statement in a test of the absorbance, A at a given
wavelength or at the maximum at about a specified wavelength
implies that the measured absorbance is within ±3% of the
stated value.
 When an assay or test prescribes the use of a reference
substance, make the spectrophotometer measurements with the
solution prepared from the reference substance by the official
directions and then with the corresponding solution prepared
from the substance being examined. Carry out the second
measurement as quickly as possible after the first, using the
same experimental conditions.

Unless otherwise specified, the requirements in the

monographs for light absorption in the tests and assays apply to
the dried or anhydrous material, where a standard is given for
loss on drying or content of water, respectively. Similar
considerations apply where standards are given for solvent
content. In calculating the result, the loss on drying or contents
of water or solvent, determined by the methods specified in the
monograph, is taken into account.

LOD (Loss on Drying

It is the loss in % w/w resulting from water and volatile

matter of any kind that can be driven off under

specified conditions. The test is carried out on

a well mixed sample of the substance. If the

 substance is the form of large crystals, reduce

the size by rapid crusting to a powder.

Method

Weigh a glass stoppered, shallow weighing bottle

that has been dried under the same conditions to be

employed in the determination. Transfer to the bottle the

quantity of the sample specified in the individual

monograph, cover it and accurately weigh the bottle and

contents. Distribute the sample as evenly as practicable

by gentle sidewise shaking to a depth not exceeding

10mm. Place the loaded bottle in the drying chamber

(over or desiccator) as directed in the monograph, remove

the stopper and leave it also in the chamber. Dry the

sample to constant weight or for the specified time and at

the temperature indicated in the monograph. After drying

is completed, open the drying chamber, close the bottle

promptly and allow it to cool to R.T. (where applicable) in

a desiccators. Weigh the bottle and contents.

Calculation

LOD (Loss on Drying)

Wt of empty bottle = xg
Wt of bottle + sample = yg
Therefore, Wt of sample = y – x g = zg
After Drying,
Wt. of bottle + sample = ag
Wt of sample = a–x
= wg

W × 100
% of LOD = ------------
z

= % w/w

Determination of Sulphated Ash

Heat a silica or platinum crucible to redness for 10

min, allow to cool in a desiccator and weigh, transfer to

the crucible 1g of the substance being examined and

weigh the crucible and the contents accurately. Ignite,

gently at first, until the substance is thoroughly charred

cool, moisten the residue with the 1 ml of H2SO4, heat

gently until the white fumes are no longer evolved and

ignite at 8000C±250C until all black particles have

disappeared. Conduct the ignition in a place protected

from air currents. Allow the crucible to cool, add a few

drops of H2SO4 and heat. Ignite as before, allow cooling

and weighing. Repeat the operation until 2 successive

weighing do not differ by more than 0.5mg.

Calculation
Sulphated Ash
Wt. of Empty Crucible = xg
Wt. of empty crucible + sample = yg
Wt. of sample taken = y–x=zg
Wt. of Crucible after ignition = wg
Sulphated Ash = w–x
= Ag

A × 100
% of Sulphated ash = ------------
Z

= B % w/w
Water Alliance
Model No. Detector 2487
Separation Module 2695

GAS CHROMATOGRAPHY

GAS CHROMATOGRAPHY
 In 1952 year James and martin invented gas liquid
chromatography and 1949-50 cremeratal fabricated the first
complete gas chromatography in their laboratory at the
University of Innsbruck Austria.

Chromatography technique is simply a method of

separation of volatile substance which is present in very minute
amount and the separation is done by adsorption and partition
process on the different adsorption column.

Gas chromatography refers to a physical process by which

a mixture is separated into its constituents by moving gas phase
over stationery adsorbent.
The gas chromatography separation occurs by partitioning
a sample between a mobile gas phase and this layer of non
volatile. Liquid coated on an inert support.

PRINCIPLE
When a gas vapors comes in contact with adsorbent a
certain amount of it get adsorbed on the solid surface according
to freundlich law

X/m = kc1/n
x = mass of gas
 m = mass of adsorbent
k = constant
c = concentration

Instrument
Carrier Gas
Helium, nitrogen, hydrogen and argon are generally used
as carrier gas. The choice of carrier gas depends upon
availability; purity and inertness towards sample and stationary
phase carrier gas flow rate generally range between 20-100
ml/min. The gas flow control is achieved by use of pressure
regulator and flow controller.
To avoid contamination carrier gas should be pure ad gas
purity can be improved by filtering it. For nitrogen as carrier gas
filtering process.

Carrier Hydro Moistur Oxygen Head

gas carbon e filter filter space
filter

This sequence prevents any hydrocarbons present in the

gas stream from reaching the oxygen filter.

Sample Injection System

To introduce a liquid sample a syringe is used. The liquid
sample is injected through a self sealing septum into head of
column and solid sample in a suitable solvent according to STP
and inject through head space.

Head Space
A head space sample is normally prepared in a vial
containing the sample, solvent, a matrix modifier and the head
space. Volatile components from complex sample mixture can
be extracted from non volatile sample components and isolated
in the head space or gas portion of a sample vial. A sample of
the gas in the head space is injected into a G-C system for
separation of all of the volatile components.
Phases of headspace vial
The gas phase is commonly referred to as the headspace
and lies above the condensed sample phase.
The sample phase contains the compound of interest. It is
usually in the form of a liquid or solid in combination with a
dilution solvent and matrix modifier.
Once the sample phase is introduced into the vial and the
vial is sealed, volatile component diffuses into the gas phase
until the head space has reached a state of equilibrium. The
sample is than taken from the headspace has reached a state of
equilibrium. The sample is then taken from the headspace.

Column
Two type of columns are used in GC:
 1.)Packed column
2.)Capillary column
Packed column are 1.5-10m in lengths and have an internal
diameter of 2-4mm. The tubing is usually made of stainless steel
or glass and contains a packing of finely divided inert, solid
support material that is coated with a liquid or solid stationary
phase. The nature of coating material determines the type of
materials will be mostly adsorbed.
Capillary columns have a very small internal diameter, on the
order of a few tenths of millimeters, and lengths between 25-
60m are common. The inner column walls are coated with the
action material. Most capillary columns are made of fused silica
with a polyimide outer coating. These columns are flexible, so a
very long column can be wound into a small coil.

Detector
Flame ionization detectors are generally used in GC.
Flame ionization detector require a flame for which the common
fuel chosen is hydrogen gas normally carrier gas used is
nitrogen gas. Both these gases are at appropriate flow rates will
enter the detector unit. Air oxygen for combustion is supplied
separately to base flame gases burning in hydrogen flame
contain very small concentration of free ions and e- resulting
from ionization. This gives an electrical conductivity which is
small. But when organic vapors enter the flamer, the
conductivity rises and this can be amplified and recorded. ECD
is also used as detector.

Computing Device:
The output from the detector is usually fed into a
potentiometer recorder or integrator cum printer plotters. These
recording devices produce a straight line when no gas/vapour is
being eluted from the column by the carrier gas
CALIBRATION

Procedure
1) Condition the column at initial oven temp.
2) In case of auto injector inject standard for six times
otherwise inject for once.
Acceptance criteria
Peak response ≥ 10. 00
RSD for area of 6 injections ≤3.0% for auto injector.

For head space sampler calibration

Adjust the chromatographic condition as per standard testing
procedure.
Procedure
1) Prepare six vials as per standard testing procedure and
inject from each 1ml gaseous phase.
2) Determine relative standard deviation of retention time
and area of peaks due to ethanol.
3) Prepare one empty vials and inject 1ml. record the
chromatogram
%age carry over = Area of ethanol in carry over injection
x100 Area of
ethanol in previous injection
Acceptance criteria
1) RSD for area of six preparations not more than 3%.
2) RSD for retention times not more than 1%.
3) % Carry over not more than 1%.

Residual solvent estimation by GC

Residual solvent in pharmaceutical are defined here as organic

volatile chemicals that are used or produced in the drug
substances during the preparation of drug product. The solvents
are not completely removed by practical manufacturing
techniques.
Residual solvents are usually organic volatile impurities that
mainly include solvents as methanol, ethanol, methylene
dichloride, toluene, etc.
Residual solvent estimation
Method of analysis:

1) Instrumentation
a) GC with flame ionization detector.
b) Head space
c) Data handling system
d) Fused silica capillary column

2) GC parameters (SET acc to STP)

 a) Oven temp. -1
b) Time -1
c) Rate
d) Injection temp.
e) Oven temp.
f) Time 2
g) FID temp.
h) Carrier gas N2 flow

3) Headspace parameter
a) Oven temp.
b) Needle temp
c) Transfer temp.
d) GC cycle temp.
e) Thermo state time
f) Pressurize time
) Thermo state time
f) Pressurize time
g) Inject time
h) Withdrawal time
i) Head space mode
j) Vial venting
k) Head space carrier pressure
l) Split
Procedure
 i) Prepare the standard as mentioned in STP of residual
solvent
ii) Note the area counts of eluting peaks form
chromatographic report.
Calculation

Product (ppm): AT x DS x P x 1000

AS DT
Where,
AT: Average area count of the peak of the reagent used
in sample solution.
AS: Average area count of the peak of the reagent used in
standard solution.
DS: Dilution factor of the standard preparation.
P: Purity in % of reagent standard.

HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

High performance liquid chromatography is a form of column

chromatography used frequently in biochemistry and analytical
chemistry. It is some times referred to as high pressure liquid
chromatography. HPLC is used to separate components of a
mixture by using a verity of chemical interaction between the
substance being analyzed and chromatography column.

Principle

HPLC enables to evaluate components of a mixture by virtue of

their differential distribution between the mobile phase and the
stationary phase, migration of solute component can only occur
when it is only dissolved in the mobile phase. Thus the solute
that have a high distribution into the stationary phase will elute
more slowly than those that distribute more readily into the
mobile phase and the two will therefore undergo
chromatographic separation.

The separation of components by chromatography depends on

the deference in their equilibrium depends on the differences in
their equilibrium distribution co efficient (K) between the
stationery and mobile phases:

K = Cs/ Cm

Where Cs and Cm are the concentration of the solute in the

stationery and mobile phases respectively solute with high K
values. Elute more slowly than those with low K value.
SYSTEM SUITABILITY

Column efficiency is defined in terms of number of theoretical

plates per meter (N) by the expression.

N = 5.54 VR/LWh2
VR = Distance along the base line between the pt. of
injection and
A ⊥ drop from maximum of peak.
L = Length of column in meters.
Wh = Width of the peak at half peak height.

Resolution factor (Russ) between the measured peaks on the

chromatogram should be greater than 1.0 and is given by

1.18 (VRb – VRS)

Rs = ----------------------
Whs + Whb

Where VRb and VRs are the distances along the base line between
the point of injection and ⊥ drawn from the maximum of two
adjacent peaks, Whs and Whb are the peak widths measured at
half peak height.
Tailing factor of peak = Wx/2A

Wx is the width of the peak of one twentieth of the peak height,

a – distance between the ⊥dropped form the peak maximum.

Apparatus

Solvent delivery system (Pump)

The pump is one of the most important component of
HPLC since is performance affect directly effect retention time,
reproducibility and detector sensitivity. The pumps deliver
steady stream of solvent from the reservoir to the detector
through the column. The pump can deliver solvent at a pressure
up to 6000 PSI.
Several types of pumps available for use with HPLC
analysis they are reciprocating piston pumps, syringe type pump
and constant pressure pumps.
The mobile phase acts as a carrier for the sample solution.
A sample solution is injected into the mobile phase of an assay
through injector part. The eluting power of mobile phase is
determined by its overall polarity, the polarity of stationary
phase and nature of the sample component.
Pumps and for quantitative analysis should be constructed
for material inert to corrosive mobile phase components and be
capable of delivering the mobile phase at a const. rate with
minimal fluctuations over extended period of time.

Injector
After dissolution in mobile phase and other suitable
solutions, compounds to be injected either manually by syringe
or loop injectors, or automatically by auto samples. The latter
consist of a carousel or rack to hold sample vials with tops that
have a pierce able Septum or Stopper and an injection device to
transfer sample from the vials to a loop from which is it loaded
into chromatograph.
A syringe can be used for manual injections of samplers
through septum when column head pressure is less than 70
atmospheres. At higher pressure an injection valve is essential.
Stop flow injections and micro volume injection sampling
valves are used.

Detector
Many compendia HPLC methods require the use of
spectrophotometric detectors such as detector consist of a flow
through cell mounted at the end of column. A beam of UV
radiation passes through the flow cell and into detector. As
compound elute from the column, they pass through the cell and
absorb the radiation, resulting in measurable energy level
changes. Fixed, variable and multiwave length detectors are
widely available.

Data collection devices

Modern data station receive and store detector output and
print out chromatographs complete with peak heights, peak
areas, and sample identification and method varieties. They are
also used to program the liquid chromatograph, controlling most
variables and providing for long periods of unattended
operations.

Columns

The columns are made from precision bore polished

stainless steel taking, typical dimensions being 10 to 30cm long
and 4 to 5mm internal diameter. The packing used in modern
HPLC columns consist of small, rigid, particle having a narrow
particle – size distribution.

Types of packing are

1. Porous, Polymeric beads
2. Porous – layer beads
3. Total porous
4. Total porous silica particles
There are various columns that are secondary to separating
column or stationary phase

1) Guard column
2) Derivatizing columns
3) Capillary columns
4) Microbore and small bore fast columns
5) Preparatory columns

Solvent

Selection of mobile phase is most important HPLC. In

general, polar materials is separated using partition
chromatography while non polar. Substances are separated using
absorption chromatograph. Better sample separation is achieved
by matching the polarities of the sample and packing and by
using a solvent that has different polarity. Beside polarity,
hydrogen bonding London dispersing forces also affect the
solvent strength.

Note : The presence of air bubbles in the mobile phase which

impair the detector single causing spike on the chromatograph
can be eliminated by degreasing the mobile phase by ultrasonic
vibrations Both single and double detector are commercially
available.
Type of analysis carried by high performance liquid
chromatography:

1.)Chromatographic purity
2.)Related substances(Impurity profile)
3.)Reaction monitoring
4.)Assay

ASSAY :

Assay is done to check the purity level of the product formed

form the substrate or the raw substrate or the raw material used
for its production. Product formed is checked for it spurity level
before it is dispatched. A particular arithmetical value it should
have for the desired parameters as per set by the STP.

Method of analysis:
1. Reagents: Different reagents are depending upon the
solubility of the product being formed.
2. Buffer solution: This solution is prepared to form the
mobile phase for the analysis system.
3. Mobile Phase: Mobile phase according to STP.
4. Chromatography System: Different chromatography
parameters are set for the sample to be analyzed.
* Instrument: HPLC equipped with UP detector.
 * Injection volume: A particular amount of the
sample is passed through the column in a particular
time and this parameter is set at very low value
mostly 20µ l.
* Run time: The analysis time is set to get the
required information form the chromatogram
produce through analysis.
REACTION MONITORING:

Production process in the pharma companies has a no. of stages.

After adding the batch to the reactor there are no. of steps it
undergoes to form the required product. Intermediate
compounds formed at each stage. It’s very important to keep a
continuous check on the intermediates formed so that we can
find out whether the reaction is going in right direction or not.
This process of continuous check is known as reaction
monitoring. Definition: Reaction monitoring is done to check
whether the reaction is going in the right direction or not i.e.
required intermediates are formed or not and also to check the
extent up to which the reaction got completed.
 HPLC
CALIBRATION OF DETECTOR 2487
CALIBRATION OF 2695 SEPARATION MODULE

Procedure
The following are to be calibrated as per standard operating
procedure.
1) Pump
2) Detector
3) Manual injector
4) Column oven

I) Pump: Pump shall be calibrated for the following

1) Flow rate accuracy
2) F low rate consistency
3) Compositional accuracy

II) Detector: Detector shall be calibrated for the following

1) Wavelength accuracy
 2) Detector linearity
3) Detector precision

III) Injector: Injector shall be calibrated for the following

1) Injector accuracy
2) Injector linearity
3) Injector precision and carousel check.
 QUALITY ASSURANCE

Quality assurance is a wide ranging concept covering all the

matter that individually or collectively influences quality of
products with regards to pharmaceuticals, quality assurance may
be divided into four major area, quality control, production,
distribution and inspection.

The assurance of product quality depends on more than just

proper sampling and adequate testing of various components
and the finished dosage form. Prime responsibility or
maintaining product quality during production rests with the
manufacturing department. Removal of responsibility from
manufacturing for producing a quality product can result in
imperfect composition, such as ingredients missing, sub potent
or super potent addition of ingredient, or mix up of ingredients:
mistake in packing and fillings, such as product contamination,
mislabeling or deficient package; and lack of conformance to
product registration. Quality assurance personal must establish
control or checkpoints to monitor the quality to the product as it
is processed and completion of manufacture. These begin with
raw materials and component testing and include in-process,
packaging, labeling and finished product testing as well as batch
Austin and stability monitoring.

Responsibility of quality assurance department :to formulate

system for the implementation of cGMP and, to ensure the
preparation, approval and implementation of standard operating
procedures, standard cleaning procedures, specifications,
standard test procedures, cleaning, validation, protocols, batch
manufacturing records, batch packaging records etc. sampling of
in process and finished products, reserve samples and stability
samples.
To review and approve validation protocols, to review changes
in product process, equipments, or any other changes as per SOP
“change control program”.
To release or reject the batch after reviewing the batch
documents to assure that the batch has been manufactured as per
the standard batch formula and there are no deviations and
deviations, if any, are record and authorized.

To conduct audits of method, results, system and process.

To release the finished products and to maintain the reserve

samples of finished product and batch documents.

To investigate market complaints and to maintain market

complaints investigation records.

To issue, control, review and retrieval of batch documents.

To investigate and disposition of incidents/OOS and approval of

deviations.

To ensure effective storage, retrieval, control and retention of all

completed documents.

Parts of quality assurance

1. documentation cell
2. In process quality control (IPQC)
3. Validation
4. Self inspection
5. Training and development
Documentation cell: the documentation cell contains all the
master formula, batch production records, and also document of
sole of company.

(a) Master formula record: master formula record for each

product should be prepared, endorsed and dated by a
competent and responsible individual. The master
formula shall include the following information’s:

1. The name of the product, a description of the dosage form,

and its strength.

2. The complete list of ingredients, designated by whole

names and codes sufficiently specific to indicate any
special characteristic.

3. The quantity by weight or volume of each ingredient,

regardless of whether it appears in the finished product.

4. The standards or specifications of each ingredients used in

the product.

5. An appropriate statement concerning any calculated excess

of an ingredient.

6. Appropriate statements of theoretic yield at various stages

and the termination of processing.

7. Manufacturing and control instructions, specifications,

precautions and special notations to be followed.

8. A detailed description of the closures, contrariness,

labeling, packaging, and other finishing material.

(b) Batch production record: batch production record

should be prepared, maintained and control for each
batch of product. The record includes date, specific
 code or identification number for each ingredient
employee, weights or measures of components and
products in the course of processing, result of in-process
and control testing, and the endorsement of the
individual performing and supervising each step of the
operation.

(c) Control of production procedures: to ensure that the

product have intended characteristics of identity
strength, quality and purity, production and the related
in-process quality control procedures should be rigidly
followed as required by the master formula record or
the batch production record. To a large extent, IPQC is
concerned with providing accurate, specific, and
definite descriptions of procedures to be employed form
the receipt of raw materials to the release of the finished
dosage forms.

In process quality control (IPQC)

The important function of the in-process quality assurance

program to ensure that the finished dosage form have uniform
purity and quality within a batch and between batches.this is
accomplished by identifying critical steps in the manufacturing
process and controlling them within defined limits.

IPQC: - do the following test to perform quality to product in

process
1) weight variation test :
2) disintegration test :
3) friability test :
4) hardness test :
5) moisture content test :
QA before start up:-

(1) environmental and microbiological control and

sanitation:-

To ensure that finished product meet high standard of quality

and purify, and effective sanitation program is required. A
successful program must be. Taken within and outside plant
to control insects and rodents. Personal cleanliness and
proper hair covering are required.

Floor, walls and cooling should be resistant to external forces

and should be easily cleaned.

Adequate ventilation, temperature and humidity and water

supply are also be checked.

QA should review of monitor the following:

Sanitation

Ventilation: - filter condition and changes, pressure gauze,

humidity and temperature
Water supply

(2) Manufacturing working formula procedure

(MWFP):
A documentation of the component material and processing step
to gather with the production operation specifications and
equipment to be used, make up the MWFP.

A working formula procedure formula should be prepared for

each batch size that is produced.

QA personal must review and check the working formula

procedures for each production batch before, during and after
production for following detail.
 (a) Signature & date if issue given by a responsible or QA
employee.

(b) Proper identification by name and dosage form, item

no, lot no, effective date of document, reference to the
superseded version, amount, lot and code no., of each
raw material utilized.

(c) Initializing of each step by two of operator involved.

(d) Starting and finishing time of each operation.

(e) Equipment to be used and etc setup.

(f) Proper labeling of released component and equipment.

RAW MATERIAL: QA should check the origin container of

released raw material for cleanliness before takes to production
department. They are weighed on environment controlled
weighing area and are transferred to secondary container which
is properly labeled and are then stored with proper
identification.

MANUFACTURING EQUIPMENT: QA personal must

ensure that manufacturing equipment is designed and located
and maintained so that it facilities through cleaning is suitable
for its intended use and minimize contamination.
QA personal should as certain that proper equipment and
tooling for manufacturing stage is being used. Equipment must
be identified by label, lot no., dosage form, item no. etc.

Quality assurance at start- up

(a)Raw material processing: - only released and properly

labeled raw material is allowed in the processing area.
 (1) QA personnel should be checked and
verify the humidity and temperature in area
are within limits or not if not then inform it
to production department.
(2) QA personnel should verify and document
the proper equipmentation, addition of
ingredient, mixing, drying and filtering
time.
(3) At certain point sample are to be taken to
the quality control lab for potency assay.

(b)Compounding:- QA personnel are responsible for as

ascertaining that all containers of raw material are properly
labeled and stage is the compounding staging area that they are
clean and the manufacturing equipment is properly identified as
to product, strength stem no.

CGMP requires that IPUA be adequately documented

throughout all stages of manufacturing.

© Packaging material control: - QA personnel can check

(1) properties of contains tightness

(2) Moisture and vapour tightness regardless of containers

construction.

(3) Physical or chemical changes of contains upon

prolonged contract with product.

(4) Toxicity of material needed for the container

construction.

(5) Compatibility between contains and product.

(d) Label control:- QA personnel inspects and verifies all

packaging component and equipment to be used for
 packaging operation to ensure that it has proper
identification that the line has been thoroughly cleared
and all the material from previous packing operation
has been completely removed.

FINISHED PRODUCT CONTROL:-

Specifications: Final testing of finished product is making in the

quality control laboratories. These tests are designed to
determine compliance with specifications. Thus, the testing of
the finished product for compliance with predetermined
standards prior to release of the product for packaging, and
subsequent distribution is a critical factor for quality assurance.
the design of test parameters, procedures, and specification is
made during product development.

Quality assurance during packaging operation: - if the

quality control dept. confirms that the product complies with the
specifications and if QA audit indicates manufacturing
operations are satisfactory the product is released to packing
department.

(a) QA personnel should periodically inspect the

packaging lines and should checked and labeled
container for compliance with the written
specifications.

(b) QA should perform an independent inspection and

select finished preservations samples at random
form each lot.

(c) It should be consisting of at beast twice the

quantity necessary to perform the entire test
required to determine.
 (g) Auditing: - GMP requires that the manufacturing
process be adequately documented throughout all stages
of the operation.

Records keeping area are

(a) Individual components, raw material packaging
materials, master formula and procedure.
(b) Batch production
(c) Packing and labeling operations
(d) Lab in process and finished control testing
(e) Proper signing and dating by at least two individual
independent for each operation.

Assurance of manufacturing practices

(1) Personnel: one criterion from a successful quality
assurance program is the encouragement of quality
consciousness in the personnel of the entire company.
Proper selection, training, and motivation of production,
packaging in the control personnel are vital to produce
quality pharmaceutical consistency. The degree to
which the desired quality of the product is attainable is
proportional to the attitudes or desires of the individual
working in production, packaging and control. It is
essential that qualified personnel be employed to
supervise the formulation, processing, sampling, testing,
packaging and labeling of the drug product, and the
competent staff be placed in charge of the maintenance
of machinery, equipment and sanitations.

(2) Equipments and Buildings: equipments and buildings

used in the manufacture, processing, packaging,
labeling, storage or control of drug should be suitable
design, size, construction and location should be
maintained in a clear and orderly manner. The building
should provide adequate space for the orderly
placement of materials and equipments to minimize any
risks of mix-ups or cross contamination.
 (3) Control of records:
Master formula records
Batch production records
Control of production procedures

(4) Packing control: at some time before the manufacture

of a product is completed, a packaging record bearing
an identification number is issued to the packaging
section. This record specifies the packaging material to
be used, operation to be performed, and the quantity to
be packaged. Simultaneously, requisitions and issue for
the product to be packaged and or the packaging and
printed materials, such as labels, containers, inserts,
brochures, cartons and the shipping cases.

VALIDATION
Validation of a process in the demonstration that controlling
the critical step of a process results in the products of
repeatable attributes (e.g., content uniformity) or causes a
reproducible event (e.g., sterilization).

The concept of applying a systems approach to

pharmaceutical manufacture and control, requiring validation
of the process and qualification of equipment, personnel, and
so forth, received considerable impetus when it was
recognized.

Types of process validation

(1) prospective validation
(2) reterospective validation
(3) concurrent validation
(4) revalidation

Types of equipments qualifications

(1) design qualifications (DQ)
(2) installation qualification (IQ)
 (3) operations qualifications (OQ)
(4) performance qualifications

PROCESS VALIDATION
It would normally be expected that process validation be
completed prior to the distribution of a finished product that is
intended for sale (prospective validation) where this is not
possible, it may be necessary to validate process during routine
production (concurrent validation) processes which have been in
use for some time without any significant changes may also be
validated according to an approved protocol (retrospective
validation).

Prospective validation: in prospective validation the validation

protocol is executed before the process is put in to commercial
use. During the product development phase the production
process should be broken down in to individual steps. Each step
should be evaluated on the basis or experience or theoretical
consideration to determine the critical parameters that may
affect the quality of finished product. A series of experiment
should be design to determine critically of these factors. Each
experiment should be planned and documented fully in an
authorized protocol.

All equipments, productions and environment and the analytical

testing methods to be used should have been fully validated.
Master batch documents can be prepared only after the critical
parameters of the process have been identified and machine
settings, components specifications and environment conditions
have been determined, by using this defined process a series of
batches should be produced. In theory, the number of the
process runs carried out and observations made should be
sufficient to allow the normal extent variations and trends to be
establish to provide sufficient data for evaluation. It is generally
considered acceptable that three consecutive batches/runs within
the final agreed parameters, giving product of the desired quality
would constitute a proper validation of the process. In practice,
it may take some considerable time to accumulate these days.
Some considerations should be exercised when selecting the
process validation strategy. Amongst these should be the use of
different lots of active raw materials and major excipients,
batches produced on different shifts, the use of different
equipments and facilities dedicated of commercial production,
operating range of critical process, and a through analysis of the
process data in case of prequalification and revalidation.
During the processing of the validation batches, extensive
sampling and testing should be performed on the product at
various stages, and should be documented comprehensively.
Detail testing should also be done on the final product in its
package.
Upon completion of the review, recommendation should be
made on the extent of monitoring and the in-process control
necessary for routine production. These should be incorporated
into the manufacturing record and packaging record or
appropriate standard operating procedures. Limits, frequencies
and action to be taken in the even to the limits being exceeded
should be specified.

Concurrent validation: in using this approach there is the always

the risk of having to modify process parameters or specifications
over a period of time. This situation often leads to question
regarding disposition of the batches that had already been
released for the sale subsequently known to have undesired
quality characteristics.
Concurrent validation may be the practical approach some
circumstance. Example:

- When a previously validated process is being

transferred to a third party contract
manufacturer or to another manufacturing
units.
 - Where the product is different strength of a
previously validated product with the same
ratio of active/inactive ingredients.

- When the number of lots evaluated under the

retrospective validation were not sufficient to
obtain a high degree assurance demonstrating
that the process is fully under control.

- When the number batches produced are

limited.

Retrospective validation:

In many establishments, processes that are stable and in routine

use have not under gone a formally documented validation
process. Historical data may be utilized to provide necessary
documentary evidence that the processes are validated.

The steps involved in this type of validation still require the

preparation of protocol, the reporting of the results of the data
review, leading to a conclusion and recommendations.

Retrospective validation is only acceptable for well established

detailed process and will be inappropriate where there have been
recent changes in the formation of the product, operating
procedures, equipments and facility.

The source of data for retrospective validation should includes

amongst others, batch documents, process control charts,
maintenance log book, process capability studies, finished
product results, including trend analysis and stability results.

For the purpose of retrospective validation studies, it is

considered acceptable that data for a minimum ten consecutive
batches produced to be utilized. When the less than ten batches
are available, it is considered that the data are not sufficient to
demonstrate retrospective that the process is fully in control. In
such cases the study should be supplemented with concurrent or
prospective validation.

Some of the essential elements for retrospective validation are:

• Batches manufactured for a defined period ( minimum of

last ten consecutive batches )

• Number of lots released per year.

• Batch size / strength / manufacture / year / period.

• Master manufacturing/packaging/documents

• Current specification for active materials/finished

products.

• List of process deviation, corrective actions and changes to

manufacturing documents.

• Data for stability testing for several batches.

• Trend analysis including those for quality related

complaints.

Process revalidation

Revalidation provides the evidence that changein a process and/

or the process environments that are introduced do not adversely
affect the process characteristics and product quality.
Documentation requirement will be the same as for the initial
validation of the process.

Revalidation becomes necessary in certain situations. Some of

the changes that require revalidation are as follows.
 • Changes in raw materials properties such as density,
viscosity, particle size, distribution , moisture etc. that
may affect the process of product.

• Changes in the source of active raw material

manufacture.

• Changes in packing material (primary container and

closure system).

• Changes in the process (such as mixing time, drying

temperature, and batch size).

• Changes in the equipment (e.g. addition of automatic

detection system). Changes of equipment which involves
the replacement of equipment on a “like for like” basis
would not requires the validation except that this new
equipment must be qualified.

• Changes in the plant/facility.

Qualification phases

Qualification of instruments is not a single, continuous process

but instead results from many discrete activities. For
convenience, these activities have been grouped in to 4 phases
of qualification. These phases are described below:

1) design qualification
2) installation qualification
3) operation qualification
4) performance qualification
DESIGN QUALIFICATION

The design qualification activity is most suitably performed by

the instrument developer/manufacturer. Since the instrument
design is already in the place for the commercial off the shelf
(COTS) systems, the user does not need to repeat all aspects of
DQ. However, users should ensure that COTS instruments are
suitable for their intended application and that the manufacture
has adopted a quality system for developing, manufacturing and
testing. Users should also establish that manufacturers and
vendors adequately support installation, services and training.
Method for ascertaining the manufacturer’s design qualification
and as instruments suitability for its intended use depend on the
nature of the instrument, the complexity of the proposed
application, and the extent of users previous interaction to the
manufacturer. Vendor audits or required vendor supplied
documentation satisfy the DQ requirement. The required scope
and comprehensiveness of the audits and documentation vary
with user’s familiarity with the instrument and their previous
interactions with the vendor.

INSTALLATION QUALIFICATION
Installation qualification is a documented collection of activities
needed to install an instrument in the users environment. IQ
applies to a new, pre-owned and an existing on-site but not
previously qualified instrument. The activities and
documentation associated with IQ are as follows:-

 system description: provide a description for the

instrument, including its manufacturer, model, serial
number, software version etc. use drawings and flowcharts
where appropriate.
 Instrument delivery: ensure that the instruments, software,
manuals, supplies and any other accessories arrive with the
instrument, manuals and documentation should be
obtained.

 Utilities/facilities/environment: verify that the installation

site satisfactorily meets vendor specified environmental
requirements. A commonsense judgment for the
environment suffices: one need not measure the exact
voltage for a standard – voltage instrument or the exact
humidity reading for an instrument that will operate at
ambient condition.

 Network and data storage: some analytical systems users

to provide network connections and data storage
capabilities at the installation site. If this is the case,
connect the instrument to the network and check its
functionality.

 Assembly and installation: assemble and install the

instrument and perform any initial diagnostics and testing.
Assembly and installation of a complex instrument are
best doneby the vendor of specialized engineer, whereas
user can assemble and install simple ones. For example
instruments, vendor establish installation tests and guide
provide valuable baseline reference for determining
instrument acceptance. Any abnormal event observed
during assembly and installation merits documenting. If
the pre-owned or unqualified existing instruments require
assembly and installation, perform the tasks as specified
here, and then perform the installation verification
procedures described below.

 Installation verification: perform the initial diagnostics and

testing of the instruments after installation. On obtaining
acceptable results, the user (when present) and the
installing engineer should confirm that the installation was
 successful before proceeding with the next qualification
phase.

Operational qualification:

After a successful IQ instrument is ready for OQ testing. The

OQ phase may consist of these test parameters:

 Fixed parameters: these test measures the instruments

nonchanging, fixed parameters such as length, height,
weight etc. if the vendor supplied specification for these
parameters satisfy the user, he or she may waive the test
requirement, however, if the user wants to confirm the
parameters, testing can be performed at the users site.
Fixed parameters do not change over the life of the
instrument and therefore never need redetermining. Note:
these tests could also be performed during the IQ phase
and, if so, fixed parameters need not tobe redetermined as
part of OQ testing.

 Secure data storage, backup, and archive: when required,

secure data handling, such as storage, back up and
archiving should b tested at the user site according to
written procedures

 Instruments function tests: test important functions to

verify that the instruments operates as intended by the
manufacturer and required by the user. The user should
select important instrument parameters for testing
according to the instruments intended use. Vendor
supplied information is useful in identifying specifications
for these parameters. Test should be designed to evaluate
the identified parameters. Users, or their qualified
designees, should perform these tests to verify that the
instrument meets vendor and users specifications.
PERFORMANCE QUALIFICATIONS

After the IQ AND OQ have been performed, the instruments

continued suitability for its intended use is proved through
performance qualification. The PQ phase includes these
parameters:

 Performance checks: set up a test or series of tests to

verify an acceptable performance of an instrument for its
intended use. PQ test are usually based on the instruments
typical on site applications. Some test may resemble those
performed during OQ, but the specifications for their
results can be set differently if required. PQ test are
performed routinely on a working instrument, not just on a
new instrument at installation. Therefore, PQ
specifications can slightly less rigorous than OQ
specifications. Nevertheless, user specification for PQ test
should evince trouble free instrument operation vis-à-vis
the intended applications. PQ test should be performed
independent of the routine analytical testing performed on
the instrument. Like OQ testing , the test can be modular
or holistic. Since many modules within a system interact,
holistic tests generally prove more effective by evaluating
the entire system and not just the system individual
modules. Testing frequency depends on the ruggedness of
the instrument and criticality of the tests performed.
Testing may be unscheduled- for example, each time of
the instrument is used. Or it may be scheduled to occur at
regular intervals; e.g., weekly, monthly and yearly.
Experience with the instrument can influence this decision.
Generally, the same PQ tests are repeated each time so that
a history of the instruments performance can be
completed. Some system suitability tests or quality control
checks that run concurrently with the test sample also
imply that the instrument is performing suitably. However,
 through system stability tests can supplement periodic PQ
tests, they cannot replace them.

Preventive maintenance and repairs: when PQ test (s) fail to

meet specifications, the instrument requires maintenance or
repair. For many instruments a periodic preventive maintenance
may also be recommended. Revelant PQ test(s) should be
repeated after the needed maintenance to repair to ensure that
the instruments remains qualified.

Standard operating procedure for operation, calibration and

maintenance: establish standard operating procedure to maintain
and calibrate the instrument. Use a log book, binder or
electronic record to document each maintenance and
calibrations activity.
REFERENCES
CONTENTS BOOK NAME PAGE NO. PUBLISHED BY
U.V. & Visible 1) Indian A76 – A77 Controller of
spectroscopy Pharmacopoeia 1996, Publication
Vol. II,

2) USPNF2006 2770 - 2774 Assian edition

HPLC 1) Indian A67 – A68 Controller of
Pharmacopoeia 1996, Publication
Vol. II,

2) USPNF2006 2644 – 2649 Assian edition

Gas Chromatography 1) Indian A65 – A67 Controller of
Pharmacopoeia 1996, Publication
Vol. II,

2) USPNF2006 2643 – 2644 Assian edition

Gas Chromatography USPNF 2006 2643 - 2644 Assian edition
Polarimeter Indian 554 – 556 Controller of
Pharmacopoeia 1996, A80 – A 82 Publication
Vol. II, A93
1. FDA, “Analytical procedures and methods validation:
chemistry, manufacturing, and controls,” federal Register
(notices) 65 (169), 52, 776-52, 777 (30 August 2000).
.
2. Amersham biosciences Inc. text on, “Affinity
chromatography: principles and methods”, 18-1022-29,
1997.

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