A Report

On
Industrial Training in
RAP Analytical Reaserch And Training centre
Second Floor, Circle Plaza, Sarda Circle, Nashik,

From 20 th NOV to 19 th Dec

Submitted by Ms. Sangale Yogita Amrutrao

To

PRES’S College Of Pharmacy (For Women) Chincholi

In partial fulfilment of the requirement of degree of

B.Pharmacy
 ACKNOWLEDGEMENT

Any accomplishment involves the guidance of many. I take

this opportunity to sincerely thank you to all those who were
involved in the successful completion of my training.

Firstly, I thank Mr. Rohan pawar under whose guidance I

have completed my training.

I would like to express our special thanks to the entire staff of

the RAP analytical Reasearch center for their invaluable help,
time to time support and for their co-operation. We are
grateful to Mr. Vikas kunde sir for providing necessary help to
get this training done.

Thanking you,
RAP Analytical Reaserch Center…
Introduction :

Name of the company: Rap Analytical Reasearch And Training

centre
Address of the company: seconf floor, circle plaza, sarda circle, Nashik,
Maharashtra 422001

Director:Mr. Rohan pawar

Owner of the company:Mr Rohan pawar

About RAP:
About RAP

'RAP' a new concept ever first time opened in Nashik,

established in 2012, which is playing a worthwhile role in the
field of Education and ultimately Pharmaceutical, Chemicals,
Bio-organics, Petroleum & many other Industries.

They observe number of students (B.Pharm., B.Sc., M.

Pharm., M.Sc., and others) who are good at theory but lack in
practical skills, lack in ideas and operation of sophisticated
analytical instruments.

When students join industries they do not have experience of

operating analytical in struments so that they spend more
time in learning the instruments and industries can't afford
to train them. With this purpose, they have started analytical
Instrument training center to train students when they ally
doing their academic course and which will also help them to
cope their needs. Through this endeavor they want to make
students able to get job in esteemed fields as these analytical
instruments has been widely Organic, Petroleum and many
more.exploited in the industries like Pharmaceutical, Food,
Chemical, Bio

RAP Analytical Research And Training Center is happy to see

we all and would like to con tribute in developing your skills
while you run towards your career success. I am glad to tell
you that RAP Analytical has trained more than 800 students
yet and almost more than 75% of them got recruited in
different pharma and other MNC's. We have also con sulted
more than 500 students for their Master's and PhD research
work. They are reached up and collaborated with many edu
cational organizations in Maharashtra.

Features
Well equipped Laboratory
Projector equipped Classroom with graceful environment
Hands on practical training on each instrument
Easy solutions for troubleshooting
Guidance for analytical method development
Enhancement in practical skills
Data analysis
Recent developments in analytical methods
Training includes :
1.Industrial guidelines and standards

1. Industry standards are a set of criteria within an industry

relating to the standard functioning and carrying out of
operations in their respective fields of production. In other
words it is the generally accepted requirements followed
by the members of an industry. It provides an orderly and
systematic formulation, adoption, or application of
standards used in a particular industry or sector of the
economy. Industry standards vary from one industry to
another
2. ROLE OF INDUSTRY STANDARD: Administration and
the legislative bodies are also benefited by the Industry
standard. They govern the practical as well as the
technological standards as per the legal requisites.
Standardization facilitates a healthy competition and
designing of new concepts. Industry standard ascertains
the rank of an industry in the economic set up of a
country. Optimum standards facilitate the creation of
political as well as business related advantages
3. The reason being that the industry standard is worked out
in consonance with the expertise of the corporate houses
and different segments of the society. In a nut shell,
industry standard is a crucial tool in acquiring industry
goals related to managerial, technological as well as
political. Therefore, setting standards for the industry
whether in the domestic market or international market
provides assurance of transparency. The ultimate aim of
setting industry standard is to provide a platform for giving
shape to new creations.
2. GMP :
Good Manufacturing Practice (GMP) is a system for ensuring
that products are consistently produced and controlled
according to quality standards. It is designed to minimize the
risks involved in any pharmaceutical production that cannot be
eliminated through testing the final product.

GMP covers all aspects of production from the starting

materials, premises, and equipment to the training and
personal hygiene of staff. Detailed written procedures are
essential for each process that could affect the quality of the
finished product. There must be systems to provide
documented proof that correct procedures are consistently
followed at each step in the manufacturing process - every time
a product is made.

3.GLP :
Good Laboratory Practice (GLP) is aquality system concerned with
the organizational process and the conditions under which a study is
planned, performed, monitored,recorded, archived and reported
Why GLP?
ƒ Development of quality test data
ƒ Mutual acceptance of data
ƒ Avoid duplication of data
ƒ Avoid technical barriers to trade
ƒ Protection of human health and the
Environment

Scope
Non-clinical safety testing of test items
contained in
Pharmaceutical products
Pesticide products
Cosmetic products
Veterinary drugs
Food and feed additives

The GLP Principles

1. Test facility organization and personnel
2. Quality Assurance (QA) program
3. Facilities
4. Apparatus, materials and reagents
5. Test systems
6. Test and reference items
7. Standard Operating Procedures (SOP’s)
8. Performance of the study
9. Reporting of study results
10. Storage and retention of records and materials5.Different
Analytical Techniques
Quality Assurance program
General
ƒ Documented Quality Assurance (QA) Program
ƒ Designated individuals as members of the QA team directly
responsible to the management
ƒ QA members not to be involved in the conduct of the study being
assured
Responsibilities of the QA Personnel
ƒ Access to the updated study plans and SOPs
ƒ Documented verification of the compliance of
study plan to the GLP principals
ƒ Inspections to determine compliance of the
study with GLP principles. Three types of
inspection
ƒ Study-based inspections
ƒ Facility-based inspections
ƒ Process-based inspections
4.Different Instrumental technique
Modules
HPLC, GC, UV-VIS,
Dissolution App., Disintegra-
tion App. & Friability App.
. Introduction
. Chromatographic Parameters
. Principles
. Working
. Instrumentation
. Data Analysis/Interpretation
. Quantitative and Qualitative Analysis
. Method Development
. Method Validation
. Troubleshooting
. Applications
. Calculations
. Practical Training

HPLC (high performance liquid chromatography)

• Introduction
• Historical perspective
• The HPLC system
• Separation criteria in HPLC
• Separation principles in HPLC
• Transport of solutes through the column
• Separation principles: selective dilution
• Selective retention
• Chromatographic parameters for characterizing separations
• Elution modes in HPLC
• Methodology and instrumentation

High Performance Liquid Chromatography (HPLC) is a form of

column chromatography that pumps a sample mixture or
analyte in a solvent (known as the mobile phase) at high
pressure through a column with chromatographic packing
material (stationary phase). The sample is carried by a moving
carrier gas stream of helium or nitrogen. HPLC has the ability to
separate, and identify compounds that are present in any
sample that can be dissolved in a liquid in trace concentrations
as low as parts per trillion. Because of this versatility, HPLC is
used in a variety of industrial and scientific applications, such
as pharmaceutical, environmental, forensics, and chemicals.

Sample retention time will vary depending on the interaction

between the stationary phase, the molecules being analyzed,
and the solvent, or solvents used. As the sample passes
through the column it interacts between the two phases at
different rate, primarily due to different polarities in the analytes.
Analytes that have the least amount of interaction with the
stationary phase or the most amount of interaction with the
mobile phase will exit the column faster.

Instrumentation:
Main components in an HPLC system include the solvent
reservoir, or multiple reservoirs, a high-pressure pump, a
column, injector system and the detector.
Detection Gas Regulator
limit recommendation recommendation
Purge gas

≤ 1 ppm HiQ Helium 5.0 BASELINE C106

series

The reservoir holds the solvent, which is referred to as the

mobile phase because it moves. There are usually a minimum
of two reservoirs in a system, with each holding up to 1000 cc
of solvent and usually fitted with a gas diffuser through which
helium can be bubbled. A pump is used to generate a specified
flow of the mobile phase. Although manual injection of samples
is still possible, most HPLCs are now fully automated and
controlled by computer. The injector, or auto sampler,
introduces the solvent into a phase stream that carries the
sample into the high pressure (up to 400 bar) column, which
contains specific packing material needed to effect separation.
The packing material is referred to as the stationary phase
because it is held in place by the column hardware.

A detector is needed to see the separated compound bands as

they elute from the high pressure column. The information is
sent from the detector to a computer which generates the
chromatogram. The mobile phase exits the detector and is
either sent to a waste, or collected, as desired.

Helium sparging is an effective method of degassing the mobile

phase to avoid unstable baselines caused by dissolved air.
Nitrogen is used as a nebulisation gas in Evaporative Light
Scattering Detector (ELSD) where the solvent is evaporated
from the sample leaving a mist as is measured.
Gas chromatography

Gas chromatography (GC) is an analytical technique used to

separate the chemical components of a sample mixture and
then detect them to determine their presence or absence
and/or how much is present. These chemical components
are usually organic molecules or gases. For GC to be
successful in their analysis, these components need to be
volatile, usually with a molecular weight below 1250 Da, and
thermally stable so they don’t degrade in the GC system. GC
is a widely used technique across most industries: for
quality control in the manufacture of many products from
cars to chemicals to pharmaceuticals; for research purposes
from the analysis of meteorites to natural products; and for
safety from environmental to food to forensics. Gas
chromatographs are frequently hyphenated to mass
spectrometers (GC-MS) to enable the identification of the
chemical components.

As the name implies, GC uses a carrier gas in the separation,

this plays the part of the mobile phase (Figure 1 (1)). The
carrier gas transports the sample molecules through the GC
system, ideally without reacting with the sample or
damaging the instrument components.

The sample is first introduced into the gas chromatograph

(GC), either with a syringe or transferred from an
autosampler (Figure 1 (2)) that may also extract the chemical
components from solid or liquid sample matrices. The
sample is injected into the GC inlet (Figure 1 (3)) through a
septum which enables the injection of the sample mixture
without losing the mobile phase. Connected to the inlet is
the analytical column (Figure 1 (4)), a long (10 – 150 m),
narrow (0.1 – 0.53 mm internal diameter) fused silica or
metal tube which contains the stationary phase coated on
the inside walls. The analytical column is held in the column
oven which is heated during the analysis to elute the less
volatile components. The outlet of the column is inserted
into the detector (Figure 1 (5)) which responds to the
chemical components eluting from the column to produce a
signal. The signal is recorded by the acquisition software on
a computer to produce a chromatogram
After injection into the GC inlet, the chemical components of
the sample mixture are first vaporized, if they aren’t already
in the gas phase. For low concentration samples the whole
vapour cloud is transferred into the analytical column by the
carrier gas in what is known as splitless mode. For high
concentration samples only a portion of the sample is
transferred to the analytical column in split mode, the
remainder is flushed from the system through the split line
to prevent overloading of the analytical column.

Once in the analytical column, the sample components are

separated by their different interactions with the stationary
phase. Therefore, when selecting the type of column to use,
the volatility and functional groups of the analytes should be
considered to match them to the stationary phase. Liquid
stationary phases mainly fall into two types: polyethylene
glycol (PEG) or polydimethylsiloxane (PDMS) based, the
latter with varying percentages of dimethyl, diphenyl or mid-
polar functional groups, for example cyanopropylphenyl.
Like separates like, therefore non-polar columns with
dimethyl or a low percentage of diphenyl are good for
separating non-polar analytes. Those molecules capable of
π-π interactions can be separated on stationary phases
containing phenyl groups. Those capable of hydrogen
bonding, for example acids and alcohols, are best separated
with PEG columns, unless they have undergone
derivatization to make them less polar.

The final step is the detection of the analyte molecules when

they elute from the column. There are many types of GC
detectors, for example: those that respond to C-H bonds like
the flame ionization detector (FID); those that respond to
specific elements for example sulfur, nitrogen or
phosphorus; and those that respond to specific properties
of the molecule, like the ability to capture an electron, as is
used with the electron capture detector (ECD).

Uv instrument

Ultraviolet–visible spectroscopy or ultraviolet–visible

spectrophotometry (UV–Vis or UV/Vis) refers to absorption
spectroscopy or reflectance spectroscopy in part of
the ultraviolet and the full, adjacent visible regions of
the electromagnetic spectrum. This means it uses light in the
visible and adjacent ranges. The absorption or reflectance in the
visible range directly affects the perceived color of the
chemicals involved. In this region of the
spectrum, atoms and molecules undergo electronic transitions.
Absorption spectroscopy is complementary to fluorescence
spectroscopy, in that fluorescence deals with transitions of
electrons from the excited state to the ground state, while
absorption measures transitions from the ground state to the
excited state.[1]
Molecules containing bonding and non-bonding electrons (n-
electrons) can absorb energy in the form of ultraviolet or visible
light to excite these electrons to higher anti-bonding molecular
orbitals.[2] The more easily excited the electrons (i.e. lower
energy gap between the HOMO and the LUMO), the longer the
wavelength of light it can absorb. There are four possible types
of transitions (π–π*, n–π*, σ–σ*, and n–σ*), and they can be
ordered as follows :σ–σ* > n–σ* > π–π* > n–π*
UV/Vis spectroscopy is routinely used in analytical chemistry for
the quantitative determination of different analytes, such
as transition metal ions, highly conjugated organic compounds,
and biological macromolecules. Spectroscopic analysis is
commonly carried out in solutions but solids and gases may
also be studied.
• Solutions of transition metal ions can be colored (i.e., absorb
visible light) because d electrons within the metal atoms can
be excited from one electronic state to another. The colour of
metal ion solutions is strongly affected by the presence of
other species, such as certain anions or ligands. For instance,
the colour of a dilute solution of copper sulfate is a very light
blue; adding ammonia intensifies the colour and changes the
wavelength of maximum absorption (λmax).
 • Organic compounds, especially those with a high degree
of conjugation, also absorb light in the UV or visible regions
of the electromagnetic spectrum. The solvents for these
determinations are often water for water-soluble
compounds, or ethanol for organic-soluble compounds.
(Organic solvents may have significant UV absorption; not
all solvents are suitable for use in UV spectroscopy. Ethanol
absorbs very weakly at most wavelengths.) Solvent polarity
and pH can affect the absorption spectrum of an organic
compound. Tyrosine, for example, increases in absorption
maxima and molar extinction coefficient when pH increases
from 6 to 13 or when solvent polarity decreases.
• While charge transfer complexes also give rise to colours,
the colours are often too intense to be used for quantitative
measurement.

Dissolution test apparatus

Inthe pharmaceutical industry, drug dissolution testing is
routinely used to provide critical in vitro drug release
information for both quality control purposes, i.e., to assess
batch-to-batch consistency of solid oral dosage forms such as
tablets, and drug development, i.e., to predict in vivo drug
release profiles. There are three typical situations where
dissolution testing plays a vital role: (i) formulation and
optimization decisions: during product development, for
products where dissolution performance is a critical quality
attribute, both the product formulation and the manufacturing
process are optimized based on achieving specific dissolution
targets. (ii) Equivalence decisions: during generic product
development, and also when implementing post-approval
process or formulation changes, similarity of in vitro dissolution
profiles between the reference product and its generic or
modified version are one of the key requirements for regulatory
approval decisions. (iii) Product compliance and release
decisions: during routine manufacturing, dissolution outcomes
are very often one of the criteria used to make product release
decisions.
The main objective of developing and evaluating an IVIVC is to
establish the dissolution test as a surrogate for human studies,
as stated by the Food and Drug Administration (FDA Analytical
data from drug dissolution testing are sufficient in many cases
to establish safety and efficacy of a drug product without in
vivo tests, following minor formulation and manufacturing
changes (Qureshi and Shabnam, 2001). Thus, the dissolution
testing which is conducted in dissolution apparatus must be
able to provide accurate and reproducible results.