Impurities in Pharmaceuticals: Sources, Classification, and Impact

PHCH 402: Analytical Quality

Control

05:

Limit tests (10 hrs)

OUTLINE

•

Presence of impurities in pharmaceuticals and

their sources.

•

Limit tests and factors considered in their design,

negative and comparison tests.

•

Classification of limit tests: limits of soluble and

insoluble matter; moisture; volatile matter;

residual solvents; nonvolatile matter. Residue on

ignition, loss on ignition, loss on drying, ash

values, limit tests for metallic and non-metallic

impurities, other specific limit tests

•

Is there any

difference between

IMPURITY and

CONTAMINANT ?

Definition of Impurities:

•

Impurities in a pharmaceutical product may be

defined as unwanted chemicals in the product

that are not the active pharmaceutical ingredient

(API) itself (or the excipients used to manufacture

it), or which develop during formulation or upon

aging of both API and formulation.

•

They are unwanted chemicals that remain within

the formulation or API in small amounts and

which can influence quality, safety and efficacy

(QSE), thereby causing serious health hazard.

In general….

•

Impurities

 are undesirable elements or

substances that occur commonly or naturally

in a substance, thereby lowering its quality or

value. Depending on its quantity, the impurity

may or may not make the substance unfit for

its intended use.

•

On the contrary, a 

contaminant

 is an external

agent that is (or gets) added to something and

usually renders it unfit for its intended use.

Sources of Impurities

•

Major sources of impurities 

may be classified

broadly into two:

     i) Synthesis related impurities



Raw materials employed



the manufacturing process



solvent



reaction vessel



stability of the final product which can be predicted

from its chemistry i.e. degradation

Sources of Impurities….

 ii) Formulation related impurities



Physical contamination



Improper storage conditions



Atmospheric contaminant



Microbial contamination



Particulate contamination



Filth

Types of Impurities

•

According to the International Conference on

Harmonisation of Technical Requirements for

Registration of Pharmaceuticals for Human Use

(ICH), 

impurities can be classified broadly into

:

     a) Organic impurities

     b) Inorganic impurities and

     c)  Residual Solvents

I. Synthesis related impurities:

A.

Organic Impurities

:

•

The composition of impurities allows one to draw

conclusions regarding the manufacturing of the

products and its adulteration. Majority of the

impurities are characteristics of the synthetic route

of the manufacturing process.

•

Since there are several possibilities of synthesizing a

     drug, it is possible that the same product of different

sources may give rise to different impurities

Synthesis related impurities:

•

In the case of drugs prepared by multi-step

synthesis, the number and the variety of

structures of organic impurities are almost

unlimited and highly dependent on the route

and reaction conditions of the syntheses and

several other factors such as the purity of the

starting material, method of isolation,

purification, conditions of storage etc.

1. Starting Materials and Intermediates:

•

Starting materials and intermediates are the

chemical building blocks used to construct the

final form of a drug substance molecule.

Unreacted starting materials and intermediates,

particularly those involved in the last a few steps

of the synthesis, can potentially survive the

synthetic and purification process and appear in

the final product as impurities

•

For example, in the synthesis of paracetamol, p-

aminophenol  could be a starting material or an

intermediate and may appear in the final

product. There is a limit test for it in bulk

paracetamol:

Production of paracetamol from

intermediate p-amino phenol:

i.

Impurities in the Starting

Materials:

•

Impurities present in the starting materials could

be present in the final product, or follow the same

reaction pathways as the starting 

material itself,

and the reaction products could carry over to the

final product as process impurities.

•

Knowledge of the impurities in starting materials

helps to identify related impurities in the final

product, and to understand the formation

mechanisms of these related process impurities

e.g

Effect of trace impurity in starting material:

•

Tolperisone

trace impurity

•

Starting material

trace impurity

ii. Degradation products during

manufacturing

•

During manufacturing of bulk drugs

degradation of end products results in the

formation of impurities. For example

Hydrochlorothiazide has a known degradation

pathway through which it degrades to the

starting material as 1,3- disulfonamide in its

synthesis:

Dihydrochlorothiazide degradation

:

          Dihydrochloro thiazide

degradation product

iii. 

By – products:

•

In synthetic organic chemistry, getting a single end –

product with 100% yield is seldom. There is always a

chance of having by-products.

•

 can be formed through variety of side reactions, such

as incomplete reaction, over-reaction, isomerization,

    dimerization, rearrangement or unwanted reactions

     between starting materials or intermediate with

chemical reagents or catalysts . For example, in the

case of paracetamol bulk production, diacetylated

paracetamol may form as a by-product :

Formation of diacetylated paracetamol

as a by-product:

iv. Organic Reagents, Ligands and

Catalysts:

•

Chemical reagents, ligands, and catalysts used in

the synthesis of a drug substance can be carried

over to the final products as trace level

impurities. For example, carbonic acid

chloromethyl tetrahydro-pyran-4-yl ester

(

CCMTHP), which is used as an alkylating agent

in the synthesis of a beta-lactam drug substance,

was observed in the final product as an impurity

•

Pyridine, a catalyst used in the course of synthesis

of mazipredone, reacts with an intermediate to

form a pyridinium impurity

v. Organic Impurities originating from

reaction solvents:

•

Impurities in the solvents can also be source

of impurities.

•

For e.g. 2-hydroxytetrahydrofuran is an

impurity in tetrahydrofuran, which is often

used as the solvent of Grignard reagents

furan:

2-hydroxytetrahydrofuran:

B: Inorganic impurities

•

Inorganic impurities derive from the

manufacturing process for the bulk drug, and

excipients. They include the following:

1. 

Reagents, ligands, and catalysts

 The chances of having these impurities are rare:

however, in some processes, these could

create a problem unless the manufacturers

take proper care during production.

2. 

Heavy metals

The main sources of heavy metals are:

a)  the water used in the processes and the

reactors (if stainless steel reactors are used),

where acidification or acid hydrolysis takes

place. These heavy metal  impurities  can

easily be avoided using demineralized water

and glass-lined reactors.

b) Excipients: Generally, excipients may contain high

levels of heavy metals such as arsenic, bismuth,

cadmium, chromium, copper, iron, lead, mercury,

nickel and sodium.

•

Sometimes they might present in the product during

processing or may leach from packing material. For

example, excipients such as hydrogenated oils and fats,

which are produced using metal catalysts, are found to

contain high concentrations of metals (platinum and

palladium). This may be due to leaching from process

equipment or storage container.

3. 

Other materials (e.g., filter aids,

charcoal etc.)

•

filters or filtering aids such as centrifuge bags

are routinely used in the bulk drugs

manufacturing plants and in many cases,

activated carbon is also used. The regular

monitoring of fibers and black particles in the

bulk drugs is essential to avoid these

contaminations.

C. 

Residual solvents

•

Residual solvents are volatile organic

chemicals used during the manufacturing

process or generated during production

•

Residual solvents are potentially undesirable

substances.

•

either modify the properties of certain

compounds or may be hazardous to human

health.

•

Also affect physicochemical properties of the

bulk drug substances such as crystallinity,

which in turn may affect the dissolution

properties, odor and colour changes in

finished products

•

In addition, solvents used in synthesis may

contain a number of impurities which can

react with chemicals used in the synthesis to

produce impurities

Water:

•

Most commonly used solvent

•

Not considered as an impurity most of the time

•

However moisture content can be very important after

packaging as moisture content may be sufficient to cause

hydrolysis

•

Drug products may also be affected by water from

environment

•

Hydrolysis due to presence of water causes chemical

instability problems

•

Water can be present even in non-aqueous formulations in

enough quantities to cause degradation

•

It can also be a major source of microbial contamination

II. FORMULATION RELATED

IMPURITIES

A.

DOSAGE FORM RELATED (EXCIPIENTS):

•

APIs are formulated with excipients (pharmaceutical aids) into

solutions, tablets, capsules, semi-solids, aerosols and Novel

Drug Delivery Systems.

•

 During formulation, excipients are added to API to render the

product elegant. They can be sometimes heterogeneous

mixtures.

i. Excipients can be a source impurities and microbial

contamination

ii. drug – excipient incompatibility may lead to undesirable

products which can affect the therapeutic efficacy of the

product eg:  See table below

Effect of Pharmaceutical Aids on

Stability of Active Ingredients

In general…

•

Liquid dosage forms may undergo both

degradation as well as microbial

contamination

•

Water content, pH of formulation,

compatibility of cations and anions, mutual

interaction of ingredients and the primary

container are the critical factors

B. METHOD RELATED IMPURITY

•

Eg. In production of parenteral dosage form of

diclofenac Na, 1-(2,6 dichlorophenyl) indoline-

2 –one is formed as an impurity when it is

terminally sterilised by autoclaving. The

formation of this derivative and NaOH occurs

due to intramolecular cyclic reaction of

diclofenac Na by autoclave conditions (123

0

C).

•

Diclofenac Na

Indolinone derivative

C.

 Environmental related impurity

1. Temperature: 

Especially

during formulation of

vitamins and antibiotics, extreme care should

be exercised to prevent them from

degradation because these classes of

compounds are heat liable. When subjected

to extreme temperature, loss of potency takes

place (for instance drying under heat)

2. Light - UV light:

•

Light is one of the means by which the

formulation degrades because of photolytic

reaction. Exposure to light is known to be

deleterious on a number of pharmaceutical

compounds. For eg. Ergometrine injection has

been reported to be unstable under tropical

conditions of light and heat. Some other drugs

that are affected are:

Drugs Affected By Light or Catalyst

3. Humidity

•

Humidity is one of the important key factors

incase of hygroscopic compounds. It is

detrimental to both bulk powder and

formulated solid dosage form. The classic

examples are ranitidine and aspirin

D. 

Impurities on Aging (storage and

transport)

1.

Mutual interaction amongst ingredients

•

Because of the labile nature of vitamins, they undergo

degradation in different dosage forms, especially liquid

•

 degradation of vitamins such as folic acid, thiamine and

cyanocobalamines does not yield toxic impurities but they

lose their potency well below compendial specifications

•

An example of mutual interaction : presence of

nicotinamide in formulation containing four vitamins

(nicotinamide, pyridoxine, riboflavin and thiamine) causes

the degradation of thiamine to a substandard level within a

one year shelf life of vitamin–B complex  injection .

2. Instability of the Product

a.

Chemical instability

•

Impurities can also arise during storage because of chemical instability of

the pharmaceutical substance. Many pharmaceutically important

substances undergo chemical decomposition when storage conditions are

inadequate.

•

 often catalyzed by light, traces of acid or alkali, traces of metallic

impurities, air oxidation, carbon dioxide and water vapours (humidity)

•

The nature of the decomposition can easily be predicted from the

knowledge of chemical properties of the substance(s)

•

All such decompositions can be minimized or avoided by using proper

storage procedures and conditions

•

The photosensitive substances should be protected from light by storing

them in darkened glass or metal containers thereby inhibiting

photochemical decomposition.

•

 Materials susceptible to oxidation by air or attack by moisture should be

stored in sealed containers

•

if necessary the air from the containers can be displaced by an inert gas

such as Nitrogen. Oxidation can also be prevented by adding suitable

antioxidants

Types of degradation

i.

Oxidation

•

Drugs which are prone to oxidation are those that

contain OH groups directly bonded to aromatic rings

eg. catechols, conjugated-dienes, heterocyclic aromatic

rings, nitroso and nitrite derivatives e.g.

Hydrocortisone, methotrexate, adinazolam,

catecholamine, (Vitamin–A) etc.

•

In pharmaceuticals, the most common form of oxidative

decomposition is auto oxidation through a free radical

chain process commonly catalysed by metals.

•

 For example, auto-oxidation of ascorbic acid studies

reveals that cupric ion  oxidises ascorbic acid rapidly to

dehydroascorbic acid and potassium cyanide. As a result,

there is a cleavage of chain due to the formation of

copper complexes.

ii. Hydrolysis

•

A reaction in which water is the reactant causing

precipitation.

•

Most well-known examples of such reactions in

pharmaceutical compounds are esters and

amides

•

Many drugs are derivatives of carboxylic acids or

contain functional groups based on that moiety

example esters, amides, lactones, lactams, imides

and carbamates, others which are susceptible to

acid-base hydrolysis include aspirin, atropine,

chloramphenicol,   barbiturates,

chlordiazepoxide, oxazepam and lincomycin etc.

iii. Decarboxylation

•

Some of the carboxylic acids such as 

p-amino

salicylic acid have shown loss of carbon dioxide

from carboxyl group when heated

•

For instance, photo reaction of rufloxacin tablet

enteric coated with cellulose acetate phthalate

(CAP) and sub-coating with calcium carbonate

cause hydrolysis of CAP liberating acetic acid,

which on reacting with calcium carbonate

produced carbon dioxide, a by-product that blew

off the cap from the bottle after cap was

loosened .

iv.  Photolysis

•

Photolytic cleavage on aging includes examples of

pharmaceutical drugs or products that are prone to

degradation on exposure to UV-light

•

During manufacturing process as solid or solution,

packaging or on storage, drugs like ergometrine,

nifedipine, nitroprusside, riboflavin and phenothiazines

are liable to photo oxidation

•

 involves generation of free radical intermediate, which

will degrade the products

•

For example, exposure of ciprofloxacin eye drop 0.3%

to UV light induces photolysis thereby resulting in the

formation of ethylene 

di-amine analogue of

ciprofloxacin

b. Physical instability

•

Pharmaceuticals may undergo changes in physical

properties during storage. There can be changes in crystal

size and shape, sedimentation, agglomeration and caking of

the suspended particles.

•

These physical changes are not always avoidable and may

result in significant changes in the physical appearance,

pharmaceutical and therapeutic effects of the product.

•

Particle size and consequently surface area is a critical

factor in determining the bioavailability of the low solubility

drug such as griseofulvin.

•

Physical changes such as sedimentation and caking in case

of multidose suspension may constitute hazard leading to

the possibility of under dosage and later to overdosage of

the drugs.

•

Similarly increase in the globule size of the injectable

emulsions on storage may lead to fat embolism.

E) Packaging material

•

Impurities result also from packaging materials i.e., containers

and closures

•

For most drugs the reactive species for impurities consists of:

      Water – hydrolysis of active ingredient.

      Small electrophiles – Aldehydes and carboxylic acid derivatives.

Peroxides – oxidize some drugs.

Metals – catalyze oxidation of drugs and their degradation pathway.

Extractable or leachables – Emerge from glass, rubber stoppers and

plastic materials, in which oxides like NO, SiO, CaO, MgO are major

components leached or extracted from glass.

•

Some examples of synthetic materials include styrene from

polystyrene, diethylhexylpthalate (DEHP) plasticizer in PVC, dioctyltin

iso octyl mercaptoacetate stabilizer for PVC, zinc stearate stabilizer in

PVC and polypropylene, bisphenol A from plastics etc.

Enantiomeric Impurities:

•

The majority of therapeutic chiral drugs used as pure

enantiomers are natural products. The high level of

enantioselectivity of their biosyntheses excludes the

possibility of the presence of enantiomeric impurities

•

In the case of synthetic chiral drugs, the racemate of

which is usually marketed, if the pure enantiomer is

administered, the antipode is considered to be an

impurity. The reason for its presence can be either

        a) the incomplete enantioselectivity of the syntheses

or b)  incomplete resolution of the enantiomers of the

racemate

•

 Although the ICH guideline excludes enantiomeric

impurities, pharmacopoeias consider them as ordinary

impurities

•

A single enantiomeric form of chiral drug is now considered as an

improved chemical entity that may offer a better pharmacological

profile and an increased therapeutic index with a more favourable

adverse reaction profile than the racemic mixture; and a lower dose

•

However, the pharmacokinetic profile of levofloxacin (S- Isomeric

form) and ofloxacin (R-isomeric form) are comparable, suggesting

the lack of advantages of single isomer in this regard

•

Typical examples of drugs containing enantiomeric impurities:

      a) Dexchlorophenarmine maleate (R enantiomer impurity allowed

NMT 0.5%)

      b) Timolol maleate (R enantiomer impurity allowed NMT 1%)

       c) Clopidogrel sulphate (R enantiomer impurity allowed NMT 1%)

•

In general, an individual API may contain all of the above-

mentioned types of organic impurities at levels varying from

negligible to significant

Pharmacopoeial Norms for the

Enantiomeric Impurities:

•

Many medicinal substances that contain one or more

chiral centres and that are already in the market have

been made available for pharmaceutical use as racemic

mixtures with little known about the biological

activities of the separate isomers and this is reflected

in the monograph in the pharmacopoeia.

•

Nevertheless, with increasing concern by regulatory

authorities for substances to be made available as

single isomers, tests for enantiomeric composition will

become more common

•

As a result the following recommendations have been

made by the BP 2001:

Chemical definition in monographs

    1) -In the case of substances containing a single chiral

centre, the descriptor ‘(RS)’ should be included at

the appropriate position in the chemical definition

of the substances to indicate a racemic mixture.

     2) - For substances containing multiple chiral centres

and comprising mixture of all possible stereomers the

term ‘all-rec’ should be used, for example Iso-aminile.

In those few substances existing as diastereomeric

mixtures, that is where in one or more centres the

stereochemistry is explicit but in other centres it is not,

each centre is defined either as the specific (R)- or (S) –

configuration , or as racemic (RS)-, respectively.

In future…

•

Tests:

       1) - when a monograph describes an

enantiomer, it will include both a test for specific

optical rotation under identification and a test

using methods such as chiral chromatography, to

control enantiomeric purity.

      2) - When only the racemic mixture is available,

the monograph for the racemic mixture will

simply specify a test for angle of rotation.

LIMIT TESTS

OUTLINE

•

Limit tests and factors considered in their

design, negative and comparison tests.

•

Classification of limit tests: limits of soluble

and insoluble matter; moisture; volatile

matter; residual solvents; nonvolatile matter.

Residue on ignition, loss on ignition, loss on

drying, ash values, limit tests for metallic and

non-metallic impurities, other specific limit

tests

Limit Tests

•

Limit tests are quantitative or semi-quantitative tests

designed to identify and control small amount of impurities

that are most likely to be present in a pharmaceutical

substance i.e. they assume no gross contamination, that

GMP has been followed.

•

They involve simple comparisons of opalescence, turbidity

or colour produced in test with that of fixed standards.

•

Since the amount of any single impurity present in an

official substance is usually small, the normal visible-

reaction-response to any test for that impurity is also quite

small.

•

it is therefore necessary and important to design the

individual test in such a manner as to avoid possible errors

in the hands of various analysts. It may be achieved by

taking into consideration the following 

 factors:

Factors considered in the design of Limit Tests:

GENERAL FACTORS:

(

a) 

Specificity of the Tests :

•

A test employed as a limit test should imply some

sort of selective reaction with the trace impurity.

(However, it has been observed that a less

specific test which limits a 

number of possible

impurities 

rather instantly has a positive edge

over the highly specific tests which limits only

one impurity.

•

Example : Contamination with Pb2+ and other

heavy metal impurities in Alum: these are

precipitated by 

thioacetamide as their respective

sulphides at pH 3.5.

 (

b) 

Sensitivity : 

The extent of sensitivity stipulated in a

limit test varies widely as per the standard laid down

by a pharmacopoeia. The sensitivity is governed by a

number of variable factors having a common objective

to yield reproducible results, for instance :

  (

i) 

Gravimetric Analysis : 

The precipitation is guided by

the concentration of the solute and of the precipitating

reagent, reaction time, reaction temperature and the

nature and amount of other substance(s) present in

solution.

  (

ii) 

Colour Tests : 

The production of visible and distinct

colouration may be achieved by ascertaining the

requisite quantities of reagents and reactants, time

period and above all the stability of the colour

produced.

 (

c) 

Personal Errors :  

personal errors must be

avoided as far as possible such as

:

(

i) 

Physical Impairment : 

A person suffering from

colour blindness may not be in a position to

assess colour-changes precisely ; or if he uses

bifocals he may not take the burette readings

accurately.

(

ii) 

Learning-Curve Syndrome : 

An analyst must

practise a new assay method employing ‘known’

    samples before making an attempt to tackle an

unknown sample, thereby minimising the scope

of personal errors.

SPECIFIC FACTORS

For Known Impurities:

•

For known impurities, several aspects are

taken into account in designing the test. These

include the nature of the impurity, its toxicity

and the levels likely to be found in routine

production. Analytical considerations such as

the response factor for the impurity and

practical issues such as availability of the

impurity as a reference material or reagent

also influence the test design

•

If a major and/or toxic impurity in a material is known

to have a significantly different response (more than

±20%) from that of the substance being examined in

the conditions of the test, the preferred manner of

limiting this impurity is to use a reference substance of

the impurity.

•

 If this is not possible, a reference solution of the

substance being examined containing a known amount

of the impurity may be used.  When neither of these

approaches is possible, a dilution of the solution of the

substance being examined may be used as a reference

solution. This approach is also commonly used in tests

where an impurity that is known (but not named

within the test) has a response within ±20% of that of

the substance being examined.

•

The response factor (

k) is a relative term, being the

response of equal weights of one substance 

relative to that

of another in the conditions described in the test.

•

In the context of a related substances test where a

response factor is quoted for an impurity, unless otherwise

stated, this is the expected response for that impurity in

relation to a response of unity for the substance being

examined. The way in which a response factor is to be used

in any subsequent calculation is stated in the monograph.

•

 Response factors of less than 0.2 or more than 5 are not

used. If the difference between the response of an impurity

and that of the substance being examined is outside these

limits, a different method of determination, such as a

different detection wavelength (λ) or a different method of

visualisation, is used.

For Unknown Impurities:

•

Unknown impurities may be limited by

reference to a dilution of the solution of the

substance being 

examined used as a reference

solution together with an open design of

statement limiting 'any' or 'any other' 

secondary

peak or spot. Such a reference solution may be

used in addition to those containing 

named

impurities (any other secondary peak/spot) or, in

some simple tests, control of unknown and

known (but unnamed) impurities may be exerted

by means of a comparison between the sample

solution and a dilution of this solution (any

secondary peak/spot).

For Formulated Preparations

•

 Many monographs for formulated preparations in the British

Pharmacopoeia also include tests for impurities. In general,

wherever possible a test for impurities based on that in the

monograph for the active ingredient is included with any necessary

modification.

•

Wider limits and/or additional controls may be required for

impurities arising on manufacture or  storage of the dosage form.

•

Tests for impurities in monographs for formulated preparation are

used to control not only degradation products but also by-products

of the synthetic route used for manufacture of the active

ingredient. It has been argued, for example in the ICH guideline

Q3B, that by-products of synthesis have been controlled already

during examination of the substance before formulation and that

further testing for these impurities is unnecessary. Clearly it would

be repetitious and wasteful of resources for tests, often complex in

nature, to be repeated routinely simply to demonstrate acceptably

low levels of impurities that could arise only during synthesis (as

opposed to degradation) of the active ingredient.

•

However, this information is available only to

those who know the detailed attributes of the

active raw material that has been used. For an

analyst who has access only to the dosage form,

the profile of synthesis-related impurities offers

one means of establishing whether or not the

dosage form has been prepared from an active

ingredient of pharmacopoeial quality. It is for this

reason that such tests are included in British

Pharmacopoeia monographs for formulated

preparations

•

Examples:

    i) Acetylsalicylic acid: Limit tests for salicylic

acid (degradation product) are specified in

both the active ingredient and the tablets

aspirin

salicylic acid

    ii) Chloramphenicol:

degradation product

Limit test for the degradation product is specified only

for the capsules

  iii) for metronidazole:

     limit test for the degradation product is only

specified for the tablets but not the active

ingredient:

        Metronidazole                                                      2-methyl-5-nitroimidazole

Limit tests involve either “negative” or

“comparison” tests:

•

Negative test: refers to absence of spot

•

Comparison test: comparison with the

response due to a known

concentration/quantity of reference impurity

Limit Tests for Related Substances

•

 It is usual to include a test for related substances in a monograph for a

medicinal substance. These may be manufacturing impurities

(intermediates or by-products) or degradation products or both.

•

When preparation of a monograph is initiated the manufacturer is

asked to provide information concerning:

      a)  the nature of such impurities,

      b) the reason for their presence,

      c)  the amounts that may be encountered in material prepared under

conditions of good pharmaceutical manufacturing practice

      d)  the manner in which proportions may vary on storage,

      e) an indication of the toxicity of any impurities in relation to that of

the substance itself.

    This is called an IMPURITY PROFILE which  involves detection  of  the

impurities using chromatography  -  either TLC, HPLC or GC (MS)

•

Where there is only one manufacturer of a

substance, pharmacopoeia limits are set in the

knowledge that the level of impurities in

production batches of the substance will have

been accepted by the registration authority

based on the toxicity studies and clinical trials

carried out before the granting of a licence.

•

Such studies and trials will have been carried out

on material with an impurity profile that is

qualitatively and quantitatively similar to that of

subsequent production batches.

•

Any subsequent changes to the manufacturing process

by the original manufacturer or the introduction of

material from another manufacturer utilising a

different route of synthesis will be subject to the need

to demonstrate essential similarity or to provide

equivalent data to the relevant registration authority.

•

In some cases a change in production or source may

give rise to impurities that are not adequately

controlled by the published pharmacopoeial

monograph. Appropriate revision of the monograph

will be carried out provided that the pharmacopoeial

authority is notified of the need and that it is supplied

with the relevant information

VARIOUS TYPES OF TESTS FOR QUANTITATIVE

DETERMINATIONS

•

In actual practice, it has been observed that

different 

official compendia 

describe a

number of detailed types of tests with a view

to obtain a constant and regular check that

might be possible to maintain the desired

degree of optimum purity both in the pure

pharmaceutical substances and the respective

dosage-forms made therefrom.

•

A number of such tests include

1)

Limits of soluble matters

2)

Limits of insoluble matters

3)

Limit of moisture, volatile matter and residual

solvent

4)

Limit of non-volatile matter, residue on ignition

and loss of ignition

5)

Limit of ash values e.g acid insoluble ash, water

soluble ash, water soluble extractives and

sulphated ash

6)

Limit test for acid radical impurities

7)

Limit test for metallic impurities etc.

Limits of soluble matter

•

In order to detect the presence of some very specific

impurities normally present in the official substances, the

limits of soluble impurities have been laid down in different

pharmacopoeias.

•

It is applied to limit soluble impurities which are completely

insoluble in a particular solvent .

•

Some typical examples include

i.

Water soluble barium salts are highly toxic and therefore,

strictly excluded from barium sulphate used for X-ray

work.

ii.

Limit of matter soluble in dilute HCl is applied to both

light and heavy kaolin by refluxing with dilute HCl,

filtering and evaporating the filtrate. NMT 0.5%

Limits of insoluble matter

•

The tests for clarity of solution offer a means of

limiting insoluble impurities in an official preparation.

•

Clarity of solutions for injection is important to ensure

complete freedom from particulate matter.

•

A special requirement for all injections and water for

injection is that the solution must be free from

insoluble matter on viewing against a black background

in upward, horizontal and inverted position.

•

Another example of limit test of insoluble matter is the

control of phenylbarbituric acid in phenobarbitone by

the requirement that 1 g shall be completely soluble in

5 ml of boiling ethanol (90 %) within 3 minutes.

Limits of Moisture, Volatile Matter and Residual

Solvents

•

A good number of pharmaceutical substances usually

absorb moisture on storage thereby causing deterioration.

Such an anomaly can be safely restricted and limited by

imposing an essential requirement for the loss in weight

(Loss on Drying) when the pharmaceutical chemical is

subjected to drying under specified conditions.

•

The quantum of heat that may be applied to the substance

varies widely as per the following norms :

     (

a) Nature of the substance

     (

b) Decomposition characteristics of the substance.

•

Various 

official compendia recommended different

temperatures and duration of drying either at atmospheric

or reduced pressure (vacuum). A few typical examples are

stated below :

•

There are 

four types of hydrates which may be

observed amongst the pharmaceutical chemicals,

namely :

(a)

Inorganic Salt Hydrates e.g., Magnesium

Sulphate (MgSO4.7H2O) ; Sodium Sulphate

(Na2SO4.

10 H2O).

(b)

Salts of Inorganic Cations and Organic Acids e.g.,

Calcium Lactate, Ferrous Gluconate.

(c)

Organic Hyrates e.g., Caffeine Hydrate,

Theophylline Hydrate.

(d)

Organic Substances e.g., Acacia, Hydroxymethyl

Cellulose.

Aquametry:

It refers to the determination of water

content titrimetrically with 

Karl Fischer Reagent

(KFR). 

This

technique has been used exclusively for

the determination of water content in a number of

pharmaceutical substances listed below:

•

Since the introduction of Gas-Liquid-Chromatography (GLC)

as an essential analytical tool, it has been judiciously exploited

as an useful alternative to KFR for not only determining water

content in pharmaceutical chemicals but also limiting specific

volatile substances present in them: eg:

•

For Determination of Water Content for  

Gonadorelin : (Limit

NMT : 7.0 % w/v)

•

For Limiting Specific Volatile Substance for Orciprenaline

Sulphate : (Limit of Water and Methanol : 6.0% w/w)

Limits of Non-Volatile Matter

•

Pharmaceutical chemicals belonging to the domain of

inorganic as well as organic substances contain readily

volatile matter for which the various 

official compendia

prescribe limits of non-volatile matter. It is 

pertinent to

mention here that the Pharmacopoeia usually makes a

clear distinction between substances that are readily

volatile and substances that are volatile upon strong

ignition, for instance :

(

a) 

Readily Volatile : e.g., Organic 

Substances

—alcohol (95%

v/v), isopropyl alcohol, chloroform,

halothane, anaesthetic

ether, chlorocresol and trichloroethylene ; and 

Inorganic

substances—

ammonia solution, hydrogen peroxide

solution, water for injection.

(

b) 

Volatile Upon Strong Ignition : e.g., hydrous wool fat

(lanolin).

Limits of Residue on Ignition

•

the limits of residue on ignition are basically applicable

to the following two categories of pharmaceutical

substances, namely :

   (

a) Those which are completely volatile when ignited

e.g., Hg.

   (

b) Those which undergo total decomposition thereby

leaving a residue with a definite composition e.g.,

calamine—a basic zinc carbonate that gives rise to ZnO

as the residue.

•

According to BP, 68.0 to 74.0% when ignited at a

temperature not lower than 900°C until, after further

ignition, two successive weighings do not differ, by

more than 0.2% of the weight of the residue.

Limits of Loss on Ignition

Limits on Ash Value

•

The ash values usually represent the inorganic

residue present in official herbal drugs and

pharmaceutical substances. These values are

categorized into 

four heads, namely :

    (

a) Ash Value (Total Ash),

    (

b) Acid-Insoluble Ash,

    (

c) Sulphated Ash, and

    (

d) Water-Soluble Ash.

Ash Value (Total Ash)

    Ash value normally designates the presence of inorganic

salts 

e.g., calcium oxalate found naturally in the 

drug, as

well as inorganic matter derived from external sources. The

official ash values are of prime importance in examination

of the purity of powdered drugs as enumerated below :

(

i) To detect and check adulteration with exhausted drugs e.g.,

ginger.

(

ii) To detect and check absence of other parts of the plant

e.g., cardamom fruit.

(

iii) To detect and check adulteration with material containing

either starch or stone cells that would 

modify the ash

values.

(

iv) To ensure the absence of an abnormal proportion of

extraneous mineral matter incorporated accidentally 

or due

to follow up treatment or due to 

modus operandi at the

time of collection e.g., soil, floor 

sweepings and sand.

•

The most common procedure recommended for 

crude

drugs is described below :

•

Procedure : Incinerate 2 to 3 g of the ground drug in a

tared platinum or silica dish at a temperature not

exceeding 450°C until free from carbon. Cool and

weigh. If a carbon-free ash cannot be obtained in this

way, exhaust the charred mass with hot water (DW),

collect the residue on an ashless filter paper, incinerate

the residue and filter paper, add the filtrate, evaporate

to dryness and ignite at a temperature not exceeding

450°C.

•

Calculate the percentage of ash with reference to the

air-dried drug.

Acid-Insoluble Ash

    The method described above for ‘

total ash’ present in

crude drugs containing calcium oxalate has certain

serious anomalies, namely :

• Offers variable results upon ashing based on the

conditions of ignition.

• Does not detect soil present in the drug efficaciously.

• The limits of excess of soil in the drug are not quite

definite.

    Hence, the treatment of the ‘

total ash’ with acid

virtually leaves silica exclusively and thus

comparatively 

forms a better test to detect and limit

excess of soil in the drug than does the ash. The

common procedure usually adopted for the

determination of ‘acid insoluble ash’ is given below :

•

Procedure : Place the ash, as described earlier, in

a crucible, add 15 ml DW and l0 ml hydrochloric

acid 

(~–11.5 N), cover with a watch-glass, boil for

10 minutes and allow to cool. Collect the

insoluble matter on an ashless filtre paper, wash

with hot DW until the filtrate is neutral, dry, ignite

to dull redness, allow to cool in a desiccator and

weigh. Repeat until the difference between two

successive weighings is not more than l mg.

•

Calculate the percentage of acid-insoluble ash

with reference to the air-dried drug.

A few typical examples are listed below 

:

Sulphated Ash

•

The estimation of ‘

sulphated ash’ is broadly employed in

the case of :

 (

a) Unorganized drugs e.g., colophony, podophyllum resin,

wool alcohols, wool fat and hydrous wool 

fat.

 (

b) Pharmaceutical substances containing inorganic impurities

e.g.,

•

Natural Origin : 

Spray-dried acacia, Frangula Bark,

Activated Charcoal

•

Organic Substances : 

Cephalexin, Lignocaine hydrochloride,

Griseofulvin, Diazoxide, Medazapam

, 

Saccharin.

•

Inorganic Substances : 

Ammonium chloride, Hydroxy urea.

•

Procedure : 

Heat a silica or platimum crucible to

redness for 30 minutes, allow to cool in a desiccator

and weigh. Place a suitable quantity of the substance

being examined, accurately weighed in the crucible,

add 2 ml of 1 M sulphuric acid and heat, first on a

waterbath and then cautiously over a flame to about

600°C. Continue heating until all black particles have

disappeared and then allow to cool. 

Add a few drops

of 1 M sulphuric acid, heat to ignition as before and

allow to cool.

 Add a few drops of a 16% solution of

ammonium carbonate, evaporate to dryness and

cautiously ignite. Cool, weigh, ignite for 15 minutes and

repeat the procedure to constant weight.

Water-Soluble Ash

•

Water-soluble ash is specifically useful in detecting such samples

which have been extracted with water.

•

A detailed procedure as per the official compendium is enumerated

below :

•

Procedure : The ash as described earlier, is boiled for 5 minutes

with 25 ml DW, collect the insoluble 

matter in a sintered-glass

crucible or on an ashless filter paper, wash with hot DW and ignite

for 15 minutes at a temperature not exceeding 450°. Subtract the

weight of the residue thus obtained from the weight of the ash.The

difference in weight represents the water-soluble ash. Now,

calculate the percentage of water-soluble ash with reference to the

air-dried drug.

•

A typical example of an official drug is that of ‘Ginger’, the water-

soluble ash of which is found to be not more than 6.0%.

•

Limit test for acid radical impurities

•

Limit test for metallic impurities

(REFER TO NOTES)